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Experiment: Identification of genes that are directly regulated by NMD in Arabidopsis

Experiment Description

NASCArrays Experiment Reference Number: NASCARRAYS-647

To understand the way NMD mediates responses it is important to distinguish between direct and indirect targets. We will accomplish this using a global analysis of mRNA stability in wild type and NMD mutant backgrounds. Direct NMD targets show enhanced stability exclusively in NMD mutants. We will use cordycepin to interrupt transcription in wild type and NMD mutant three week old seedlings and analyse samples at time intervals following treatment. We will take timepoints at 0hrs, 1hr, 3hrs and 6hrs. We will analyse global mRNA stability in seedlings of four of our NMD mutant lines (upf1-5, UPF2 (6-11E5), upf3-1 and smg7-1) and wild type. This experiment will generate a list of mRNAs that are directly destabilised by NMD, which can be compared and analysed for features that make them targets of NMD. Three crucial questions will be addressed by this experiment. Firstly, the mRNA decay data will allow us to identify the genes that are directly targeted by each subset of the NMD pathway. For example, targets that are independent of UPF3 will appear to have altered stability only in the upf1 and smg7 mutant backgrounds. Secondly, it will allow us to test the association of specific NMD trigger signals with subsets of NMD. Finally, the list of direct targets will facilitate identification and dissection of the downstream regulated processes.

About the Experimenter

Name:Dr Samantha Rayson
Head of Lab Name:Dr Brendan Davies
Lab:
Address:Centre for Plant Sciences, Institute of Integrative and Comparative Biology, Faculty of Biological Sciences, University of Leeds
Leeds, West Yorkshire, LS2 9JT, UNITED KINGDOM
Postcode: LS2 9JT
Country: UNITED KINGDOM
 
Telephone Number: 011334332838

All of the data available in this website/database is free, and you are free to do whatever you please with it. If you intend to publish work based on any of this data, please acknowledge us, contact the experimenter above, and either acknowledge them or use them as co-authors in the work.

About this Experiment

Experiment Type: compound_based_treatment
Number of Slides:20
 
Experimental Parameters:
parametercompound_based_treatment
Quality Control Measures Taken:
References:
 
Other Information:

Slides in this Experiment

Hybridisation Set: Rayson: Identification of genes that are directly regulated by NMD in Arabidopsis

Slide: Rayson_647-9_upf1-5_NMD_0hr_cordycepin_Rep1_ATH1

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Summary BioSource 1 Information Extraction / Labelling Protocol for Source 1 Information Slide Information Hybridisation Analysis Information for "Affymetrix MAS 5.0 Scaling Protocol" normalised data.

Summary Information

This slide has had 1 extract(s) hybridised to it (1-colour).

  • Extract 1 ("Rayson_647-9_upf1-5_NMD_0hr_cordycepin_Rep1_ATH1") was made from "Arabidopsis thaliana" and labelled using "Biotin Labelled cRNA using Affymetrix Enzo Kit".

The slide used was "Affymetrix ATH1 Arabidopsis Genome Array".

Click on the options above for more information about this slide and its preparation.

Hybridisation Protocol Information

Name of Protocol: Affymetrix Antibody Amplification Protocol
Solution:
BufferEDTA, NaCl, Tween20 according to Affymetrix protocol
Water
Blocking Agent:
Herring Sperm DNA(Source: Promega)100pg/ml
Acetylated BSA(Source: Sigma-Aldrich)0.1mg/ml
Wash Procedure:
Step 1(Source: Affymetrix)10 cycles of 2 mixes/cycle with Non-Stringent buffer
Step 2(Source: Affymetrix)4 cycles of 15 mixes/cycle with Stringent buffer
Step 3(Source: Affymetrix)Stain the probe array for 10 minutes in Streptavaidin Phycoerythrin (SAPE) solution at 25 C
Step 4(Source: Affymetrix)10 cycles of 4 mixes/cycle with Non-Stringent buffer
Step 5(Source: Affymetrix)Stain the probe array for 10 minutes in antibody solution
Step 6(Source: Affymetrix)Stain the probe array for 10 minutes in SAPE solution at 25 C
Step 7(Source: Affymetrix)15 cycles of 4mixes/cycle with Non-Stringent buffer at 30 C
Step 8(Source: Affymetrix)Hold at 25 C
Quantity Used:
Amount15 microgram Fragmented cRNA
Time: 16 hours
Concentration: 50 pg/ml
Volume: 200 microlitres
Temperature: 45 C
Other Information:
Spike(Source: Affymetrix)Control Oligo B2 to 50pM for landing lights
Spike(Source: Affymetrix)20X Eukaryotic Hybridisation Controls (bioB (1.5pM, bioC 5pM, bioD 25pM, cre 100pM)
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

BioSource 1 Information

BioSource Name: Rayson_647-9_upf1-5_NMD_0hr_cordycepin_Rep1_ATH1
Organism: Arabidopsis thaliana
Alias: upf1-5
Stock Code: N9902
Genetic Background: Col-0
Age: 3 weeks
Growth Conditions:
StratificationImbibed in the dark at 4 °C for 3 days.
ProtocolPlants were grown in a Sanyo MLR-352 H with additive humidity under a typical short day cycle (8hr light:16 hr dark) The temperature was maintained at 21-23°C and the humidity was maintained at 65 % RH. Stratified seeds were sown onto SHL compost at a density of 4 seed per cell (24 cell tray). Trays were covered in a propagator lid, which was removed 7 days after sowing. Compost was kept moist throughout.
Soil ConstituentsSinclair Potting Growth Medium (www.william-sinclair.co.uk)
Plant Spacing1 per cell
waterCompost was kept moist at all times
Temperature22 °C average, 23 °C day, 21 °C night
Humidity65 % average, 65 % day, 65 % night
MediumNone
Lighting(Source: Cool white fluorescent manufactured by SANYO. Intensity: 175µEinsteins. Wavelength: White)
Developmental Stage:
Growth Stage(Source: Paradigm Genetics)
Tissue: Aerial tissues
in vivo Treatment: Time = 0 Aerial tissues were harvested into incubation buffer (IB: 1 mM Pipes, 1 mM Sodium citrate, 1 mM Potassium chloride, 15 mM Sucrose). After 30 minutes in IB, all except the time (T) = 0 samples were transferred to IB with cordycepin (150 µg ml-1). The T = 0 samples were transferred to fresh IB without cordycepin. All flasks (including the T = 0) were transferred to a bell jar and vacuum infiltrated. Immediately post-infiltration was classed as T =0.
Additional Organism Information:
Sample DescriptionSoil-grown plant grown under short-day conditions.
Other Information:
Timecourse start procedureThe time-course started immediately after the vacuum infiltration.

Protocols for BioSource 1

Extraction Protocol Information

Name of Protocol: Qiagen / Trizol
Method:
ProtocolQiagen RNAeasy kit protocol
Type: total RNA
Amplification: Amplified during labelling process - see labelling protocol
Other Information:
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

Labelling Protocol Information

Name of Protocol: Biotin Labelled cRNA using Affymetrix Enzo Kit
Amount Labelled: Unknown
Label Name: Biotin
Labelling Method: In-vitro transcription method
Other Information:
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

Slide Information

Date Hybridised:

Array Design Information

Slide Design Name: Affymetrix ATH1 Arabidopsis Genome Array
Array Type: Affymetrix Gene Chip
Number of Spots on the Array: 22810

Scanning Protocol Information

Name: Affymetrix MAS 5.0 Standard Scanning
Width of Image Produced (Pixels): 4733
Height of Image Produced (Pixels): 4733
X Resolution (microns): 3
Y Resolution (microns): 3
Scanner Name: Agilent 2500A GeneArray Scanner
Software Name: Affymetrix Micro Array Suite 5.0

Image Analysis Information

Scaling Factor:0.744065
Name: Affymetrix MAS 5.0 Standard Image Analysis
Software Name: Affymetrix Microarray Analysis Suite 5.0

Normalisation Information

Strategy Name: Affymetrix MAS 5.0 Scaling Protocol
Algorithm: The top 2% and bottom 2% of signal intensities are excluded, then the mean is calculated. The original signal values are scaled such that the mean is made equal to 100.
Control Elements:
none
Normalisation Protocol Parameters:
Spike_AFFX-r2-P1-cre_3-5-ratio1.259097
Spike_AFFX-r2-Ec-bioB_3-5-ratio2.168324
Spike_AFFX-r2-Bs-dap_5_signal26.115332
NoiseAvg:3.08,Std:0.09,Min:2.9,Max:3.4
Control DirectionAntisense
Spike_AFFX-r2-Ec-bioB_5_detectionA
Spike_AFFX-r2-Ec-bioB_3_detectionM
Spike_AFFX-r2-Bs-thr_5_signal8.795153
#P13116
Spike_AFFX-r2-Bs-phe_M_signal16.236235
Corner-Avg:11704,Count:32
Spike_AFFX-r2-P1-cre_3_detectionP
Spike_AFFX-r2-Ec-bioB_M_signal15.784777
Spike_AFFX-r2-Ec-bioB_3_signal18.224543
Spike_AFFX-r2-Bs-lys_M_signal11.420631
Spike_AFFX-r2-P1-cre_3_signal6446.535156
Spike_AFFX-r2-Bs-lys_3-5-ratio1.416652
Spike_AFFX-r2-Bs-dap_M_signal65.262718
Spike_AFFX-r2-Bs-thr_M_detectionM
Probe Pair Threshold1
Spike_AFFX-r2-Bs-thr_3-5-ratio6.725444
Spike_AFFX-r2-Ec-bioB_avg-signal14.138073
Spike_AFFX-r2-Bs-thr_avg-signal30.360769
Spike_AFFX-r2-Bs-thr_5_detectionP
Spike_AFFX-r2-Bs-dap_3_detectionP
Spike_AFFX-r2-Ec-bioD_5_signal1417.219238
Spike_AFFX-r2-Bs-phe_5_signal8.456137
Spike_AFFX-r2-P1-cre_5_detectionP
Spike_AFFX-r2-Ec-bioD_3_signal2062.933594
RawQ2.887360
Spike_AFFX-r2-Bs-lys_5_signal15.322198
Signal(A)5.358262
%A40.618149
Signal(All)148.297058
Corner+Avg:192,Count:32
Spike_AFFX-r2-Ec-bioD_5_detectionP
Spike_AFFX-r2-Ec-bioC_3_signal380.449371
Spike_AFFX-r2-Bs-lys_3_detectionP
Spike_AFFX-r2-Bs-phe_3_signal21.618320
Spike_AFFX-r2-Ec-bioD_avg-signal1740.076416
Spike_AFFX-r2-Bs-dap_5_detectionP
Spike_AFFX-r2-Bs-phe_5_detectionA
%P57.501095
Spike_AFFX-r2-Bs-lys_avg-signal16.149683
Spike_AFFX-r2-P1-cre_avg-signal5783.250000
Spike_AFFX-r2-Ec-bioB_M_detectionP
Spike_AFFX-r2-Bs-lys_M_detectionP
#Probe Sets Exceeding Probe Pair Threshold22810
%M1.880754
Spike_AFFX-r2-Bs-thr_3_signal59.151310
Spike_AFFX-r2-Ec-bioC_5_detectionP
Spike_AFFX-r2-Bs-phe_3_detectionP
Spike_AFFX-r2-Ec-bioC_3_detectionP
Spike_AFFX-r2-Bs-phe_avg-signal15.436897
Spike_AFFX-r2-Bs-dap_3_signal136.562317
Spike_AFFX-r2-Ec-bioD_3-5-ratio1.455621
Spike_AFFX-r2-Ec-bioD_3_detectionP
Spike_AFFX-r2-Ec-bioC_5_signal353.633698
#M429
Spike_AFFX-r2-Bs-lys_5_detectionP
Spike_AFFX-r2-Bs-lys_3_signal21.706221
Spike_AFFX-r2-Bs-dap_M_detectionP
Spike_AFFX-r2-Ec-bioC_3-5-ratio1.075829
Spike_AFFX-r2-Bs-thr_M_signal23.135841
Signal(P)253.490891
Spike_AFFX-r2-Bs-phe_3-5-ratio2.556525
Spike_AFFX-r2-Bs-thr_3_detectionP
Central-Avg:10008,Count:9
Spike_AFFX-r2-P1-cre_5_signal5119.965332
Spike_AFFX-r2-Bs-phe_M_detectionP
#A9265
Signal(M)19.172808
BackgroundAvg:71.08,Std:1.15,Min:69.2,Max:75.1
Spike_AFFX-r2-Ec-bioC_avg-signal367.041534
Spike_AFFX-r2-Bs-dap_avg-signal75.980125
Spike_AFFX-r2-Bs-dap_3-5-ratio5.229201
Spike_AFFX-r2-Ec-bioB_5_signal8.404900
ScaleMaskAll
BG2
VZ4
TGT100
Alpha10.040000
Alpha20.060000
SF0.744065
NF1.000000
HZ4
Tau0.015000

Slide: Rayson_647-8_smg7-1_NMD_6hr_cordycepin_Rep1_ATH1

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Get the data!

Summary BioSource 1 Information Extraction / Labelling Protocol for Source 1 Information Slide Information Hybridisation Analysis Information for "Affymetrix MAS 5.0 Scaling Protocol" normalised data.

Summary Information

This slide has had 1 extract(s) hybridised to it (1-colour).

  • Extract 1 ("Rayson_647-8_smg7-1_NMD_6hr_cordycepin_Rep1_ATH1") was made from "Arabidopsis thaliana" and labelled using "Biotin Labelled cRNA using Affymetrix Enzo Kit".

The slide used was "Affymetrix ATH1 Arabidopsis Genome Array".

Click on the options above for more information about this slide and its preparation.

Hybridisation Protocol Information

Name of Protocol: Affymetrix Antibody Amplification Protocol
Solution:
BufferEDTA, NaCl, Tween20 according to Affymetrix protocol
Water
Blocking Agent:
Herring Sperm DNA(Source: Promega)100pg/ml
Acetylated BSA(Source: Sigma-Aldrich)0.1mg/ml
Wash Procedure:
Step 1(Source: Affymetrix)10 cycles of 2 mixes/cycle with Non-Stringent buffer
Step 2(Source: Affymetrix)4 cycles of 15 mixes/cycle with Stringent buffer
Step 3(Source: Affymetrix)Stain the probe array for 10 minutes in Streptavaidin Phycoerythrin (SAPE) solution at 25 C
Step 4(Source: Affymetrix)10 cycles of 4 mixes/cycle with Non-Stringent buffer
Step 5(Source: Affymetrix)Stain the probe array for 10 minutes in antibody solution
Step 6(Source: Affymetrix)Stain the probe array for 10 minutes in SAPE solution at 25 C
Step 7(Source: Affymetrix)15 cycles of 4mixes/cycle with Non-Stringent buffer at 30 C
Step 8(Source: Affymetrix)Hold at 25 C
Quantity Used:
Amount15 microgram Fragmented cRNA
Time: 16 hours
Concentration: 50 pg/ml
Volume: 200 microlitres
Temperature: 45 C
Other Information:
Spike(Source: Affymetrix)Control Oligo B2 to 50pM for landing lights
Spike(Source: Affymetrix)20X Eukaryotic Hybridisation Controls (bioB (1.5pM, bioC 5pM, bioD 25pM, cre 100pM)
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

BioSource 1 Information

BioSource Name: Rayson_647-8_smg7-1_NMD_6hr_cordycepin_Rep1_ATH1
Organism: Arabidopsis thaliana
Alias: SALK_073354 (AF) (AN)
Stock Code: N573354
Genetic Background: Col-0 (Columbia, N60000)
Age: 3 weeks
Growth Conditions:
StratificationImbibed in the dark at 4 °C for 3 days.
ProtocolPlants were grown in a Sanyo MLR-352 H with additive humidity under a typical short day cycle (8hr light:16 hr dark) The temperature was maintained at 21-23°C and the humidity was maintained at 65 % RH. Stratified seeds were sown onto SHL compost at a density of 4 seed per cell (24 cell tray). Trays were covered in a propagator lid, which was removed 7 days after sowing. Compost was kept moist throughout.
Soil ConstituentsSinclair Potting Growth Medium (www.william-sinclair.co.uk)
Plant Spacing1 per cell
waterCompost was kept moist at all times
Temperature22 °C average, 23 °C day, 21 °C night
Humidity65 % average, 65 % day, 65 % night
MediumNone
Lighting(Source: Cool white fluorescent manufactured by SANYO. Intensity: 175µEinsteins. Wavelength: White)
Developmental Stage:
Growth Stage(Source: Paradigm Genetics)
Genetic Variation: T-DNA Insertion
Tissue: Aerial tissues
in vivo Treatment: Time = 6 hours Aerial tissues were harvested into incubation buffer (IB: 1 mM Pipes, 1 mM Sodium citrate, 1 mM Potassium chloride, 15 mM Sucrose). After 30 minutes in IB, all except the time (T) = 0 samples were transferred to IB with cordycepin (150 µg ml-1). The T = 0 samples were transferred to fresh IB without cordycepin. All flasks (including the T = 0) were transferred to a bell jar and vacuum infiltrated. Immediately post-infiltration was classed as T =0. One leaf was taken for genotyping 1 week prior to treatment. Only confirmed homozygotes were used.
Additional Organism Information:
Sample DescriptionSoil-grown plant grown under short-day conditions.
Other Information:
Timecourse start procedureThe time-course started immediately after the vacuum infiltration.

Protocols for BioSource 1

Extraction Protocol Information

Name of Protocol: Qiagen / Trizol
Method:
ProtocolQiagen RNAeasy kit protocol
Type: total RNA
Amplification: Amplified during labelling process - see labelling protocol
Other Information:
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

Labelling Protocol Information

Name of Protocol: Biotin Labelled cRNA using Affymetrix Enzo Kit
Amount Labelled: Unknown
Label Name: Biotin
Labelling Method: In-vitro transcription method
Other Information:
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

Slide Information

Date Hybridised:

Array Design Information

Slide Design Name: Affymetrix ATH1 Arabidopsis Genome Array
Array Type: Affymetrix Gene Chip
Number of Spots on the Array: 22810

Scanning Protocol Information

Name: Affymetrix MAS 5.0 Standard Scanning
Width of Image Produced (Pixels): 4733
Height of Image Produced (Pixels): 4733
X Resolution (microns): 3
Y Resolution (microns): 3
Scanner Name: Agilent 2500A GeneArray Scanner
Software Name: Affymetrix Micro Array Suite 5.0

Image Analysis Information

Scaling Factor:1.0838
Name: Affymetrix MAS 5.0 Standard Image Analysis
Software Name: Affymetrix Microarray Analysis Suite 5.0

Normalisation Information

Strategy Name: Affymetrix MAS 5.0 Scaling Protocol
Algorithm: The top 2% and bottom 2% of signal intensities are excluded, then the mean is calculated. The original signal values are scaled such that the mean is made equal to 100.
Control Elements:
none
Normalisation Protocol Parameters:
Spike_AFFX-r2-P1-cre_3-5-ratio1.147566
Spike_AFFX-r2-Ec-bioB_3-5-ratio1.064910
Spike_AFFX-r2-Bs-dap_5_signal65.832611
NoiseAvg:3.07,Std:0.11,Min:2.8,Max:3.6
Control DirectionAntisense
Spike_AFFX-r2-Ec-bioB_5_detectionA
Spike_AFFX-r2-Ec-bioB_3_detectionP
Spike_AFFX-r2-Bs-thr_5_signal39.261711
#P11644
Spike_AFFX-r2-Bs-phe_M_signal70.870392
Corner-Avg:11400,Count:32
Spike_AFFX-r2-P1-cre_3_detectionP
Spike_AFFX-r2-Ec-bioB_M_signal29.820629
Spike_AFFX-r2-Ec-bioB_3_signal25.808784
Spike_AFFX-r2-Bs-lys_M_signal43.541328
Spike_AFFX-r2-P1-cre_3_signal8184.566406
Spike_AFFX-r2-Bs-lys_3-5-ratio1.156671
Spike_AFFX-r2-Bs-dap_M_signal353.302795
Spike_AFFX-r2-Bs-thr_M_detectionP
Probe Pair Threshold1
Spike_AFFX-r2-Bs-thr_3-5-ratio6.737235
Spike_AFFX-r2-Ec-bioB_avg-signal26.621683
Spike_AFFX-r2-Bs-thr_avg-signal141.371445
Spike_AFFX-r2-Bs-thr_5_detectionP
Spike_AFFX-r2-Bs-dap_3_detectionP
Spike_AFFX-r2-Ec-bioD_5_signal2015.173096
Spike_AFFX-r2-Bs-phe_5_signal47.786598
Spike_AFFX-r2-P1-cre_5_detectionP
Spike_AFFX-r2-Ec-bioD_3_signal2802.508057
RawQ2.727270
Spike_AFFX-r2-Bs-lys_5_signal42.964600
Signal(A)8.812604
%A46.444542
Signal(All)159.765045
Corner+Avg:199,Count:32
Spike_AFFX-r2-Ec-bioD_5_detectionP
Spike_AFFX-r2-Ec-bioC_3_signal598.235168
Spike_AFFX-r2-Bs-lys_3_detectionP
Spike_AFFX-r2-Bs-phe_3_signal109.432449
Spike_AFFX-r2-Ec-bioD_avg-signal2408.840576
Spike_AFFX-r2-Bs-dap_5_detectionP
Spike_AFFX-r2-Bs-phe_5_detectionP
%P51.047787
Spike_AFFX-r2-Bs-lys_avg-signal45.400616
Spike_AFFX-r2-P1-cre_avg-signal7658.338379
Spike_AFFX-r2-Ec-bioB_M_detectionP
Spike_AFFX-r2-Bs-lys_M_detectionP
#Probe Sets Exceeding Probe Pair Threshold22810
%M2.507672
Spike_AFFX-r2-Bs-thr_3_signal264.515381
Spike_AFFX-r2-Ec-bioC_5_detectionP
Spike_AFFX-r2-Bs-phe_3_detectionP
Spike_AFFX-r2-Ec-bioC_3_detectionP
Spike_AFFX-r2-Bs-phe_avg-signal76.029816
Spike_AFFX-r2-Bs-dap_3_signal628.562500
Spike_AFFX-r2-Ec-bioD_3-5-ratio1.390703
Spike_AFFX-r2-Ec-bioD_3_detectionP
Spike_AFFX-r2-Ec-bioC_5_signal498.317657
#M572
Spike_AFFX-r2-Bs-lys_5_detectionP
Spike_AFFX-r2-Bs-lys_3_signal49.695923
Spike_AFFX-r2-Bs-dap_M_detectionP
Spike_AFFX-r2-Ec-bioC_3-5-ratio1.200510
Spike_AFFX-r2-Bs-thr_M_signal120.337234
Signal(P)303.579742
Spike_AFFX-r2-Bs-phe_3-5-ratio2.290024
Spike_AFFX-r2-Bs-thr_3_detectionP
Central-Avg:9956,Count:9
Spike_AFFX-r2-P1-cre_5_signal7132.110352
Spike_AFFX-r2-Bs-phe_M_detectionP
#A10594
Signal(M)27.967424
BackgroundAvg:70.09,Std:0.48,Min:69.1,Max:71.8
Spike_AFFX-r2-Ec-bioC_avg-signal548.276428
Spike_AFFX-r2-Bs-dap_avg-signal349.232635
Spike_AFFX-r2-Bs-dap_3-5-ratio9.547890
Spike_AFFX-r2-Ec-bioB_5_signal24.235640
ScaleMaskAll
BG2
VZ4
TGT100
Alpha10.040000
Alpha20.060000
SF1.083800
NF1.000000
HZ4
Tau0.015000

Slide: Rayson_647-7_smg7-1_NMD_3hr_cordycepin_Rep1_ATH1

Add to Slide Selection

Get the data!

Summary BioSource 1 Information Extraction / Labelling Protocol for Source 1 Information Slide Information Hybridisation Analysis Information for "Affymetrix MAS 5.0 Scaling Protocol" normalised data.

Summary Information

This slide has had 1 extract(s) hybridised to it (1-colour).

  • Extract 1 ("Rayson_647-7_smg7-1_NMD_3hr_cordycepin_Rep1_ATH1") was made from "Arabidopsis thaliana" and labelled using "Biotin Labelled cRNA using Affymetrix Enzo Kit".

The slide used was "Affymetrix ATH1 Arabidopsis Genome Array".

Click on the options above for more information about this slide and its preparation.

Hybridisation Protocol Information

Name of Protocol: Affymetrix Antibody Amplification Protocol
Solution:
BufferEDTA, NaCl, Tween20 according to Affymetrix protocol
Water
Blocking Agent:
Herring Sperm DNA(Source: Promega)100pg/ml
Acetylated BSA(Source: Sigma-Aldrich)0.1mg/ml
Wash Procedure:
Step 1(Source: Affymetrix)10 cycles of 2 mixes/cycle with Non-Stringent buffer
Step 2(Source: Affymetrix)4 cycles of 15 mixes/cycle with Stringent buffer
Step 3(Source: Affymetrix)Stain the probe array for 10 minutes in Streptavaidin Phycoerythrin (SAPE) solution at 25 C
Step 4(Source: Affymetrix)10 cycles of 4 mixes/cycle with Non-Stringent buffer
Step 5(Source: Affymetrix)Stain the probe array for 10 minutes in antibody solution
Step 6(Source: Affymetrix)Stain the probe array for 10 minutes in SAPE solution at 25 C
Step 7(Source: Affymetrix)15 cycles of 4mixes/cycle with Non-Stringent buffer at 30 C
Step 8(Source: Affymetrix)Hold at 25 C
Quantity Used:
Amount15 microgram Fragmented cRNA
Time: 16 hours
Concentration: 50 pg/ml
Volume: 200 microlitres
Temperature: 45 C
Other Information:
Spike(Source: Affymetrix)Control Oligo B2 to 50pM for landing lights
Spike(Source: Affymetrix)20X Eukaryotic Hybridisation Controls (bioB (1.5pM, bioC 5pM, bioD 25pM, cre 100pM)
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

BioSource 1 Information

BioSource Name: Rayson_647-7_smg7-1_NMD_3hr_cordycepin_Rep1_ATH1
Organism: Arabidopsis thaliana
Alias: SALK_073354 (AF) (AN)
Stock Code: N573354
Genetic Background: Col-0 (Columbia, N60000)
Age: 3 weeks
Growth Conditions:
StratificationImbibed in the dark at 4 °C for 3 days.
ProtocolPlants were grown in a Sanyo MLR-352 H with additive humidity under a typical short day cycle (8hr light:16 hr dark) The temperature was maintained at 21-23°C and the humidity was maintained at 65 % RH. Stratified seeds were sown onto SHL compost at a density of 4 seed per cell (24 cell tray). Trays were covered in a propagator lid, which was removed 7 days after sowing. Compost was kept moist throughout.
Soil ConstituentsSinclair Potting Growth Medium (www.william-sinclair.co.uk)
Plant Spacing1 per cell
waterCompost was kept moist at all times
Temperature22 °C average, 23 °C day, 21 °C night
Humidity65 % average, 65 % day, 65 % night
MediumNone
Lighting(Source: Cool white fluorescent manufactured by SANYO. Intensity: 175µEinsteins. Wavelength: White)
Developmental Stage:
Growth Stage(Source: Paradigm Genetics)
Genetic Variation: T-DNA Insertion
Tissue: Aerial tissues
in vivo Treatment: Time = 3 hours Aerial tissues were harvested into incubation buffer (IB: 1 mM Pipes, 1 mM Sodium citrate, 1 mM Potassium chloride, 15 mM Sucrose). After 30 minutes in IB, all except the time (T) = 0 samples were transferred to IB with cordycepin (150 µg ml-1). The T = 0 samples were transferred to fresh IB without cordycepin. All flasks (including the T = 0) were transferred to a bell jar and vacuum infiltrated. Immediately post-infiltration was classed as T =0. One leaf was taken for genotyping 1 week prior to treatment. Only confirmed homozygotes were used.
Additional Organism Information:
Sample DescriptionSoil-grown plant grown under short-day conditions.
Other Information:
Timecourse start procedureThe time-course started immediately after the vacuum infiltration.

Protocols for BioSource 1

Extraction Protocol Information

Name of Protocol: Qiagen / Trizol
Method:
ProtocolQiagen RNAeasy kit protocol
Type: total RNA
Amplification: Amplified during labelling process - see labelling protocol
Other Information:
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

Labelling Protocol Information

Name of Protocol: Biotin Labelled cRNA using Affymetrix Enzo Kit
Amount Labelled: Unknown
Label Name: Biotin
Labelling Method: In-vitro transcription method
Other Information:
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

Slide Information

Date Hybridised:

Array Design Information

Slide Design Name: Affymetrix ATH1 Arabidopsis Genome Array
Array Type: Affymetrix Gene Chip
Number of Spots on the Array: 22810

Scanning Protocol Information

Name: Affymetrix MAS 5.0 Standard Scanning
Width of Image Produced (Pixels): 4733
Height of Image Produced (Pixels): 4733
X Resolution (microns): 3
Y Resolution (microns): 3
Scanner Name: Agilent 2500A GeneArray Scanner
Software Name: Affymetrix Micro Array Suite 5.0

Image Analysis Information

Scaling Factor:0.818647
Name: Affymetrix MAS 5.0 Standard Image Analysis
Software Name: Affymetrix Microarray Analysis Suite 5.0

Normalisation Information

Strategy Name: Affymetrix MAS 5.0 Scaling Protocol
Algorithm: The top 2% and bottom 2% of signal intensities are excluded, then the mean is calculated. The original signal values are scaled such that the mean is made equal to 100.
Control Elements:
none
Normalisation Protocol Parameters:
Spike_AFFX-r2-P1-cre_3-5-ratio1.359704
Spike_AFFX-r2-Ec-bioB_3-5-ratio1.184198
Spike_AFFX-r2-Bs-dap_5_signal65.000885
NoiseAvg:3.05,Std:0.08,Min:2.9,Max:3.3
Control DirectionAntisense
Spike_AFFX-r2-Ec-bioB_5_detectionP
Spike_AFFX-r2-Ec-bioB_3_detectionP
Spike_AFFX-r2-Bs-thr_5_signal27.612104
#P12348
Spike_AFFX-r2-Bs-phe_M_signal49.209312
Corner-Avg:13625,Count:32
Spike_AFFX-r2-P1-cre_3_detectionP
Spike_AFFX-r2-Ec-bioB_M_signal25.922699
Spike_AFFX-r2-Ec-bioB_3_signal23.398003
Spike_AFFX-r2-Bs-lys_M_signal38.210503
Spike_AFFX-r2-P1-cre_3_signal7902.238281
Spike_AFFX-r2-Bs-lys_3-5-ratio1.699042
Spike_AFFX-r2-Bs-dap_M_signal234.752716
Spike_AFFX-r2-Bs-thr_M_detectionP
Probe Pair Threshold1
Spike_AFFX-r2-Bs-thr_3-5-ratio6.482245
Spike_AFFX-r2-Ec-bioB_avg-signal23.026405
Spike_AFFX-r2-Bs-thr_avg-signal95.449791
Spike_AFFX-r2-Bs-thr_5_detectionP
Spike_AFFX-r2-Bs-dap_3_detectionP
Spike_AFFX-r2-Ec-bioD_5_signal1854.504517
Spike_AFFX-r2-Bs-phe_5_signal39.274231
Spike_AFFX-r2-P1-cre_5_detectionP
Spike_AFFX-r2-Ec-bioD_3_signal2759.079834
RawQ2.825605
Spike_AFFX-r2-Bs-lys_5_signal30.634190
Signal(A)6.957839
%A43.586147
Signal(All)157.657654
Corner+Avg:244,Count:32
Spike_AFFX-r2-Ec-bioD_5_detectionP
Spike_AFFX-r2-Ec-bioC_3_signal583.260559
Spike_AFFX-r2-Bs-lys_3_detectionP
Spike_AFFX-r2-Bs-phe_3_signal82.090729
Spike_AFFX-r2-Ec-bioD_avg-signal2306.792236
Spike_AFFX-r2-Bs-dap_5_detectionP
Spike_AFFX-r2-Bs-phe_5_detectionP
%P54.134151
Spike_AFFX-r2-Bs-lys_avg-signal40.297821
Spike_AFFX-r2-P1-cre_avg-signal6856.986328
Spike_AFFX-r2-Ec-bioB_M_detectionP
Spike_AFFX-r2-Bs-lys_M_detectionP
#Probe Sets Exceeding Probe Pair Threshold22810
%M2.279702
Spike_AFFX-r2-Bs-thr_3_signal178.988434
Spike_AFFX-r2-Ec-bioC_5_detectionP
Spike_AFFX-r2-Bs-phe_3_detectionP
Spike_AFFX-r2-Ec-bioC_3_detectionP
Spike_AFFX-r2-Bs-phe_avg-signal56.858093
Spike_AFFX-r2-Bs-dap_3_signal462.789001
Spike_AFFX-r2-Ec-bioD_3-5-ratio1.487772
Spike_AFFX-r2-Ec-bioD_3_detectionP
Spike_AFFX-r2-Ec-bioC_5_signal544.235779
#M520
Spike_AFFX-r2-Bs-lys_5_detectionP
Spike_AFFX-r2-Bs-lys_3_signal52.048767
Spike_AFFX-r2-Bs-dap_M_detectionP
Spike_AFFX-r2-Ec-bioC_3-5-ratio1.071706
Spike_AFFX-r2-Bs-thr_M_signal79.748848
Signal(P)284.636169
Spike_AFFX-r2-Bs-phe_3-5-ratio2.090193
Spike_AFFX-r2-Bs-thr_3_detectionP
Central-Avg:12508,Count:9
Spike_AFFX-r2-P1-cre_5_signal5811.733887
Spike_AFFX-r2-Bs-phe_M_detectionP
#A9942
Signal(M)23.670923
BackgroundAvg:69.90,Std:0.39,Min:69.0,Max:70.9
Spike_AFFX-r2-Ec-bioC_avg-signal563.748169
Spike_AFFX-r2-Bs-dap_avg-signal254.180862
Spike_AFFX-r2-Bs-dap_3-5-ratio7.119734
Spike_AFFX-r2-Ec-bioB_5_signal19.758516
ScaleMaskAll
BG2
VZ4
TGT100
Alpha10.040000
Alpha20.060000
SF0.818647
NF1.000000
HZ4
Tau0.015000

Slide: Rayson_647-6_smg7-1_NMD_1hr_cordycepin_Rep1_ATH1

Add to Slide Selection

Get the data!

Summary BioSource 1 Information Extraction / Labelling Protocol for Source 1 Information Slide Information Hybridisation Analysis Information for "Affymetrix MAS 5.0 Scaling Protocol" normalised data.

Summary Information

This slide has had 1 extract(s) hybridised to it (1-colour).

  • Extract 1 ("Rayson_647-6_smg7-1_NMD_1hr_cordycepin_Rep1_ATH1") was made from "Arabidopsis thaliana" and labelled using "Biotin Labelled cRNA using Affymetrix Enzo Kit".

The slide used was "Affymetrix ATH1 Arabidopsis Genome Array".

Click on the options above for more information about this slide and its preparation.

Hybridisation Protocol Information

Name of Protocol: Affymetrix Antibody Amplification Protocol
Solution:
BufferEDTA, NaCl, Tween20 according to Affymetrix protocol
Water
Blocking Agent:
Herring Sperm DNA(Source: Promega)100pg/ml
Acetylated BSA(Source: Sigma-Aldrich)0.1mg/ml
Wash Procedure:
Step 1(Source: Affymetrix)10 cycles of 2 mixes/cycle with Non-Stringent buffer
Step 2(Source: Affymetrix)4 cycles of 15 mixes/cycle with Stringent buffer
Step 3(Source: Affymetrix)Stain the probe array for 10 minutes in Streptavaidin Phycoerythrin (SAPE) solution at 25 C
Step 4(Source: Affymetrix)10 cycles of 4 mixes/cycle with Non-Stringent buffer
Step 5(Source: Affymetrix)Stain the probe array for 10 minutes in antibody solution
Step 6(Source: Affymetrix)Stain the probe array for 10 minutes in SAPE solution at 25 C
Step 7(Source: Affymetrix)15 cycles of 4mixes/cycle with Non-Stringent buffer at 30 C
Step 8(Source: Affymetrix)Hold at 25 C
Quantity Used:
Amount15 microgram Fragmented cRNA
Time: 16 hours
Concentration: 50 pg/ml
Volume: 200 microlitres
Temperature: 45 C
Other Information:
Spike(Source: Affymetrix)Control Oligo B2 to 50pM for landing lights
Spike(Source: Affymetrix)20X Eukaryotic Hybridisation Controls (bioB (1.5pM, bioC 5pM, bioD 25pM, cre 100pM)
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

BioSource 1 Information

BioSource Name: Rayson_647-6_smg7-1_NMD_1hr_cordycepin_Rep1_ATH1
Organism: Arabidopsis thaliana
Alias: SALK_073354 (AF) (AN)
Stock Code: N573354
Genetic Background: Col-0 (Columbia, N60000)
Age: 3 weeks
Growth Conditions:
StratificationImbibed in the dark at 4 °C for 3 days.
ProtocolPlants were grown in a Sanyo MLR-352 H with additive humidity under a typical short day cycle (8hr light:16 hr dark) The temperature was maintained at 21-23°C and the humidity was maintained at 65 % RH. Stratified seeds were sown onto SHL compost at a density of 4 seed per cell (24 cell tray). Trays were covered in a propagator lid, which was removed 7 days after sowing. Compost was kept moist throughout.
Soil ConstituentsSinclair Potting Growth Medium (www.william-sinclair.co.uk)
Plant Spacing1 per cell
waterCompost was kept moist at all times
Temperature22 °C average, 23 °C day, 21 °C night
Humidity65 % average, 65 % day, 65 % night
MediumNone
Lighting(Source: Cool white fluorescent manufactured by SANYO. Intensity: 175µEinsteins. Wavelength: White)
Developmental Stage:
Growth Stage(Source: Paradigm Genetics)
Genetic Variation: T-DNA Insertion
Tissue: Aerial tissues
in vivo Treatment: Time = 1hour Aerial tissues were harvested into incubation buffer (IB: 1 mM Pipes, 1 mM Sodium citrate, 1 mM Potassium chloride, 15 mM Sucrose). After 30 minutes in IB, all except the time (T) = 0 samples were transferred to IB with cordycepin (150 µg ml-1). The T = 0 samples were transferred to fresh IB without cordycepin. All flasks (including the T = 0) were transferred to a bell jar and vacuum infiltrated. Immediately post-infiltration was classed as T =0. One leaf was taken for genotyping 1 week prior to treatment. Only confirmed homozygotes were used.
Additional Organism Information:
Sample DescriptionSoil-grown plant grown under short-day conditions.
Other Information:
Timecourse start procedureThe time-course started immediately after the vacuum infiltration.

Protocols for BioSource 1

Extraction Protocol Information

Name of Protocol: Qiagen / Trizol
Method:
ProtocolQiagen RNAeasy kit protocol
Type: total RNA
Amplification: Amplified during labelling process - see labelling protocol
Other Information:
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

Labelling Protocol Information

Name of Protocol: Biotin Labelled cRNA using Affymetrix Enzo Kit
Amount Labelled: Unknown
Label Name: Biotin
Labelling Method: In-vitro transcription method
Other Information:
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

Slide Information

Date Hybridised:

Array Design Information

Slide Design Name: Affymetrix ATH1 Arabidopsis Genome Array
Array Type: Affymetrix Gene Chip
Number of Spots on the Array: 22810

Scanning Protocol Information

Name: Affymetrix MAS 5.0 Standard Scanning
Width of Image Produced (Pixels): 4733
Height of Image Produced (Pixels): 4733
X Resolution (microns): 3
Y Resolution (microns): 3
Scanner Name: Agilent 2500A GeneArray Scanner
Software Name: Affymetrix Micro Array Suite 5.0

Image Analysis Information

Scaling Factor:0.650479
Name: Affymetrix MAS 5.0 Standard Image Analysis
Software Name: Affymetrix Microarray Analysis Suite 5.0

Normalisation Information

Strategy Name: Affymetrix MAS 5.0 Scaling Protocol
Algorithm: The top 2% and bottom 2% of signal intensities are excluded, then the mean is calculated. The original signal values are scaled such that the mean is made equal to 100.
Control Elements:
none
Normalisation Protocol Parameters:
Spike_AFFX-r2-P1-cre_3-5-ratio1.278636
Spike_AFFX-r2-Ec-bioB_3-5-ratio1.566182
Spike_AFFX-r2-Bs-dap_5_signal39.262093
NoiseAvg:3.29,Std:0.08,Min:3.1,Max:3.5
Control DirectionAntisense
Spike_AFFX-r2-Ec-bioB_5_detectionA
Spike_AFFX-r2-Ec-bioB_3_detectionP
Spike_AFFX-r2-Bs-thr_5_signal17.224758
#P13366
Spike_AFFX-r2-Bs-phe_M_signal33.602783
Corner-Avg:13403,Count:32
Spike_AFFX-r2-P1-cre_3_detectionP
Spike_AFFX-r2-Ec-bioB_M_signal11.309068
Spike_AFFX-r2-Ec-bioB_3_signal16.602606
Spike_AFFX-r2-Bs-lys_M_signal16.604523
Spike_AFFX-r2-P1-cre_3_signal5859.621582
Spike_AFFX-r2-Bs-lys_3-5-ratio1.616941
Spike_AFFX-r2-Bs-dap_M_signal168.381805
Spike_AFFX-r2-Bs-thr_M_detectionP
Probe Pair Threshold1
Spike_AFFX-r2-Bs-thr_3-5-ratio5.994919
Spike_AFFX-r2-Ec-bioB_avg-signal12.837453
Spike_AFFX-r2-Bs-thr_avg-signal58.575977
Spike_AFFX-r2-Bs-thr_5_detectionP
Spike_AFFX-r2-Bs-dap_3_detectionP
Spike_AFFX-r2-Ec-bioD_5_signal1277.494263
Spike_AFFX-r2-Bs-phe_5_signal19.019075
Spike_AFFX-r2-P1-cre_5_detectionP
Spike_AFFX-r2-Ec-bioD_3_signal1906.244751
RawQ2.947079
Spike_AFFX-r2-Bs-lys_5_signal23.783770
Signal(A)5.305082
%A39.324856
Signal(All)150.500549
Corner+Avg:240,Count:32
Spike_AFFX-r2-Ec-bioD_5_detectionP
Spike_AFFX-r2-Ec-bioC_3_signal373.143219
Spike_AFFX-r2-Bs-lys_3_detectionP
Spike_AFFX-r2-Bs-phe_3_signal50.344212
Spike_AFFX-r2-Ec-bioD_avg-signal1591.869507
Spike_AFFX-r2-Bs-dap_5_detectionP
Spike_AFFX-r2-Bs-phe_5_detectionP
%P58.597107
Spike_AFFX-r2-Bs-lys_avg-signal26.281752
Spike_AFFX-r2-P1-cre_avg-signal5221.166992
Spike_AFFX-r2-Ec-bioB_M_detectionP
Spike_AFFX-r2-Bs-lys_M_detectionP
#Probe Sets Exceeding Probe Pair Threshold22810
%M2.078036
Spike_AFFX-r2-Bs-thr_3_signal103.261024
Spike_AFFX-r2-Ec-bioC_5_detectionP
Spike_AFFX-r2-Bs-phe_3_detectionP
Spike_AFFX-r2-Ec-bioC_3_detectionP
Spike_AFFX-r2-Bs-phe_avg-signal34.322021
Spike_AFFX-r2-Bs-dap_3_signal286.404602
Spike_AFFX-r2-Ec-bioD_3-5-ratio1.492175
Spike_AFFX-r2-Ec-bioD_3_detectionP
Spike_AFFX-r2-Ec-bioC_5_signal325.997803
#M474
Spike_AFFX-r2-Bs-lys_5_detectionP
Spike_AFFX-r2-Bs-lys_3_signal38.456959
Spike_AFFX-r2-Bs-dap_M_detectionP
Spike_AFFX-r2-Ec-bioC_3-5-ratio1.144619
Spike_AFFX-r2-Bs-thr_M_signal55.242149
Signal(P)252.597275
Spike_AFFX-r2-Bs-phe_3-5-ratio2.647038
Spike_AFFX-r2-Bs-thr_3_detectionP
Central-Avg:11634,Count:9
Spike_AFFX-r2-P1-cre_5_signal4582.712891
Spike_AFFX-r2-Bs-phe_M_detectionM
#A8970
Signal(M)19.231604
BackgroundAvg:74.06,Std:0.48,Min:72.9,Max:75.1
Spike_AFFX-r2-Ec-bioC_avg-signal349.570496
Spike_AFFX-r2-Bs-dap_avg-signal164.682831
Spike_AFFX-r2-Bs-dap_3-5-ratio7.294685
Spike_AFFX-r2-Ec-bioB_5_signal10.600686
ScaleMaskAll
BG2
VZ4
TGT100
Alpha10.040000
Alpha20.060000
SF0.650479
NF1.000000
HZ4
Tau0.015000

Slide: Rayson_647-5_smg7-1_NMD_0hr_cordycepin_Rep1_ATH1

Add to Slide Selection

Get the data!

Summary BioSource 1 Information Extraction / Labelling Protocol for Source 1 Information Slide Information Hybridisation Analysis Information for "Affymetrix MAS 5.0 Scaling Protocol" normalised data.

Summary Information

This slide has had 1 extract(s) hybridised to it (1-colour).

  • Extract 1 ("Rayson_647-5_smg7-1_NMD_0hr_cordycepin_Rep1_ATH1") was made from "Arabidopsis thaliana" and labelled using "Biotin Labelled cRNA using Affymetrix Enzo Kit".

The slide used was "Affymetrix ATH1 Arabidopsis Genome Array".

Click on the options above for more information about this slide and its preparation.

Hybridisation Protocol Information

Name of Protocol: Affymetrix Antibody Amplification Protocol
Solution:
BufferEDTA, NaCl, Tween20 according to Affymetrix protocol
Water
Blocking Agent:
Herring Sperm DNA(Source: Promega)100pg/ml
Acetylated BSA(Source: Sigma-Aldrich)0.1mg/ml
Wash Procedure:
Step 1(Source: Affymetrix)10 cycles of 2 mixes/cycle with Non-Stringent buffer
Step 2(Source: Affymetrix)4 cycles of 15 mixes/cycle with Stringent buffer
Step 3(Source: Affymetrix)Stain the probe array for 10 minutes in Streptavaidin Phycoerythrin (SAPE) solution at 25 C
Step 4(Source: Affymetrix)10 cycles of 4 mixes/cycle with Non-Stringent buffer
Step 5(Source: Affymetrix)Stain the probe array for 10 minutes in antibody solution
Step 6(Source: Affymetrix)Stain the probe array for 10 minutes in SAPE solution at 25 C
Step 7(Source: Affymetrix)15 cycles of 4mixes/cycle with Non-Stringent buffer at 30 C
Step 8(Source: Affymetrix)Hold at 25 C
Quantity Used:
Amount15 microgram Fragmented cRNA
Time: 16 hours
Concentration: 50 pg/ml
Volume: 200 microlitres
Temperature: 45 C
Other Information:
Spike(Source: Affymetrix)Control Oligo B2 to 50pM for landing lights
Spike(Source: Affymetrix)20X Eukaryotic Hybridisation Controls (bioB (1.5pM, bioC 5pM, bioD 25pM, cre 100pM)
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

BioSource 1 Information

BioSource Name: Rayson_647-5_smg7-1_NMD_0hr_cordycepin_Rep1_ATH1
Organism: Arabidopsis thaliana
Alias: SALK_073354 (AF) (AN)
Stock Code: N573354
Genetic Background: Col-0 (Columbia, N60000)
Age: 3 weeks
Growth Conditions:
StratificationImbibed in the dark at 4 °C for 3 days.
ProtocolPlants were grown in a Sanyo MLR-352 H with additive humidity under a typical short day cycle (8hr light:16 hr dark) The temperature was maintained at 21-23°C and the humidity was maintained at 65 % RH. Stratified seeds were sown onto SHL compost at a density of 4 seed per cell (24 cell tray). Trays were covered in a propagator lid, which was removed 7 days after sowing. Compost was kept moist throughout.
Soil ConstituentsSinclair Potting Growth Medium (www.william-sinclair.co.uk)
Plant Spacing1 per cell
waterCompost was kept moist at all times
Temperature22 °C average, 23 °C day, 21 °C night
Humidity65 % average, 65 % day, 65 % night
MediumNone
Lighting(Source: Cool white fluorescent manufactured by SANYO. Intensity: 175µEinsteins. Wavelength: White)
Developmental Stage:
Growth Stage(Source: Paradigm Genetics)
Genetic Variation: T-DNA Insertion
Tissue: Aerial tissues
in vivo Treatment: Time = 0 Aerial tissues were harvested into incubation buffer (IB: 1 mM Pipes, 1 mM Sodium citrate, 1 mM Potassium chloride, 15 mM Sucrose). After 30 minutes in IB, all except the time (T) = 0 samples were transferred to IB with cordycepin (150 µg ml-1). The T = 0 samples were transferred to fresh IB without cordycepin. All flasks (including the T = 0) were transferred to a bell jar and vacuum infiltrated. Immediately post-infiltration was classed as T =0. One leaf was taken for genotyping 1 week prior to treatment. Only confirmed homozygotes were used.
Additional Organism Information:
Sample DescriptionSoil-grown plant grown under short-day conditions.
Other Information:
Timecourse start procedureThe time-course started immediately after the vacuum infiltration.

Protocols for BioSource 1

Extraction Protocol Information

Name of Protocol: Qiagen / Trizol
Method:
ProtocolQiagen RNAeasy kit protocol
Type: total RNA
Amplification: Amplified during labelling process - see labelling protocol
Other Information:
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

Labelling Protocol Information

Name of Protocol: Biotin Labelled cRNA using Affymetrix Enzo Kit
Amount Labelled: Unknown
Label Name: Biotin
Labelling Method: In-vitro transcription method
Other Information:
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

Slide Information

Date Hybridised:

Array Design Information

Slide Design Name: Affymetrix ATH1 Arabidopsis Genome Array
Array Type: Affymetrix Gene Chip
Number of Spots on the Array: 22810

Scanning Protocol Information

Name: Affymetrix MAS 5.0 Standard Scanning
Width of Image Produced (Pixels): 4733
Height of Image Produced (Pixels): 4733
X Resolution (microns): 3
Y Resolution (microns): 3
Scanner Name: Agilent 2500A GeneArray Scanner
Software Name: Affymetrix Micro Array Suite 5.0

Image Analysis Information

Scaling Factor:0.679695
Name: Affymetrix MAS 5.0 Standard Image Analysis
Software Name: Affymetrix Microarray Analysis Suite 5.0

Normalisation Information

Strategy Name: Affymetrix MAS 5.0 Scaling Protocol
Algorithm: The top 2% and bottom 2% of signal intensities are excluded, then the mean is calculated. The original signal values are scaled such that the mean is made equal to 100.
Control Elements:
none
Normalisation Protocol Parameters:
Spike_AFFX-r2-P1-cre_3-5-ratio1.174601
Spike_AFFX-r2-Ec-bioB_3-5-ratio1.618227
Spike_AFFX-r2-Bs-dap_5_signal24.440525
NoiseAvg:3.29,Std:0.39,Min:2.9,Max:5.6
Control DirectionAntisense
Spike_AFFX-r2-Ec-bioB_5_detectionP
Spike_AFFX-r2-Ec-bioB_3_detectionP
Spike_AFFX-r2-Bs-thr_5_signal11.722185
#P13768
Spike_AFFX-r2-Bs-phe_M_signal24.577280
Corner-Avg:12223,Count:32
Spike_AFFX-r2-P1-cre_3_detectionP
Spike_AFFX-r2-Ec-bioB_M_signal10.914367
Spike_AFFX-r2-Ec-bioB_3_signal24.774134
Spike_AFFX-r2-Bs-lys_M_signal25.097504
Spike_AFFX-r2-P1-cre_3_signal6029.170410
Spike_AFFX-r2-Bs-lys_3-5-ratio1.707117
Spike_AFFX-r2-Bs-dap_M_signal159.423477
Spike_AFFX-r2-Bs-thr_M_detectionP
Probe Pair Threshold1
Spike_AFFX-r2-Bs-thr_3-5-ratio8.536723
Spike_AFFX-r2-Ec-bioB_avg-signal16.999310
Spike_AFFX-r2-Bs-thr_avg-signal53.352612
Spike_AFFX-r2-Bs-thr_5_detectionP
Spike_AFFX-r2-Bs-dap_3_detectionP
Spike_AFFX-r2-Ec-bioD_5_signal1501.824341
Spike_AFFX-r2-Bs-phe_5_signal22.379297
Spike_AFFX-r2-P1-cre_5_detectionP
Spike_AFFX-r2-Ec-bioD_3_signal1975.232910
RawQ2.882504
Spike_AFFX-r2-Bs-lys_5_signal21.519581
Signal(A)4.560712
%A37.974571
Signal(All)143.996902
Corner+Avg:204,Count:32
Spike_AFFX-r2-Ec-bioD_5_detectionP
Spike_AFFX-r2-Ec-bioC_3_signal417.484131
Spike_AFFX-r2-Bs-lys_3_detectionP
Spike_AFFX-r2-Bs-phe_3_signal47.134800
Spike_AFFX-r2-Ec-bioD_avg-signal1738.528564
Spike_AFFX-r2-Bs-dap_5_detectionP
Spike_AFFX-r2-Bs-phe_5_detectionP
%P60.359493
Spike_AFFX-r2-Bs-lys_avg-signal27.784506
Spike_AFFX-r2-P1-cre_avg-signal5581.060547
Spike_AFFX-r2-Ec-bioB_M_detectionP
Spike_AFFX-r2-Bs-lys_M_detectionP
#Probe Sets Exceeding Probe Pair Threshold22810
%M1.665936
Spike_AFFX-r2-Bs-thr_3_signal100.069046
Spike_AFFX-r2-Ec-bioC_5_detectionP
Spike_AFFX-r2-Bs-phe_3_detectionP
Spike_AFFX-r2-Ec-bioC_3_detectionP
Spike_AFFX-r2-Bs-phe_avg-signal31.363792
Spike_AFFX-r2-Bs-dap_3_signal290.766541
Spike_AFFX-r2-Ec-bioD_3-5-ratio1.315222
Spike_AFFX-r2-Ec-bioD_3_detectionP
Spike_AFFX-r2-Ec-bioC_5_signal399.342285
#M380
Spike_AFFX-r2-Bs-lys_5_detectionP
Spike_AFFX-r2-Bs-lys_3_signal36.736435
Spike_AFFX-r2-Bs-dap_M_detectionP
Spike_AFFX-r2-Ec-bioC_3-5-ratio1.045429
Spike_AFFX-r2-Bs-thr_M_signal48.266605
Signal(P)235.218750
Spike_AFFX-r2-Bs-phe_3-5-ratio2.106179
Spike_AFFX-r2-Bs-thr_3_detectionP
Central-Avg:9455,Count:9
Spike_AFFX-r2-P1-cre_5_signal5132.950684
Spike_AFFX-r2-Bs-phe_M_detectionP
#A8662
Signal(M)17.296869
BackgroundAvg:72.97,Std:0.68,Min:70.1,Max:74.3
Spike_AFFX-r2-Ec-bioC_avg-signal408.413208
Spike_AFFX-r2-Bs-dap_avg-signal158.210175
Spike_AFFX-r2-Bs-dap_3-5-ratio11.896902
Spike_AFFX-r2-Ec-bioB_5_signal15.309430
ScaleMaskAll
BG2
VZ4
TGT100
Alpha10.040000
Alpha20.060000
SF0.679695
NF1.000000
HZ4
Tau0.015000

Slide: Rayson_647-4_WT_6hr_cordycepin_Rep1_ATH1

Add to Slide Selection

Get the data!

Summary BioSource 1 Information Extraction / Labelling Protocol for Source 1 Information Slide Information Hybridisation Analysis Information for "Affymetrix MAS 5.0 Scaling Protocol" normalised data.

Summary Information

This slide has had 1 extract(s) hybridised to it (1-colour).

  • Extract 1 ("Rayson_647-4_WT_6hr_cordycepin_Rep1_ATH1") was made from "Arabidopsis thaliana" and labelled using "Biotin Labelled cRNA using Affymetrix Enzo Kit".

The slide used was "Affymetrix ATH1 Arabidopsis Genome Array".

Click on the options above for more information about this slide and its preparation.

Hybridisation Protocol Information

Name of Protocol: Affymetrix Antibody Amplification Protocol
Solution:
BufferEDTA, NaCl, Tween20 according to Affymetrix protocol
Water
Blocking Agent:
Herring Sperm DNA(Source: Promega)100pg/ml
Acetylated BSA(Source: Sigma-Aldrich)0.1mg/ml
Wash Procedure:
Step 1(Source: Affymetrix)10 cycles of 2 mixes/cycle with Non-Stringent buffer
Step 2(Source: Affymetrix)4 cycles of 15 mixes/cycle with Stringent buffer
Step 3(Source: Affymetrix)Stain the probe array for 10 minutes in Streptavaidin Phycoerythrin (SAPE) solution at 25 C
Step 4(Source: Affymetrix)10 cycles of 4 mixes/cycle with Non-Stringent buffer
Step 5(Source: Affymetrix)Stain the probe array for 10 minutes in antibody solution
Step 6(Source: Affymetrix)Stain the probe array for 10 minutes in SAPE solution at 25 C
Step 7(Source: Affymetrix)15 cycles of 4mixes/cycle with Non-Stringent buffer at 30 C
Step 8(Source: Affymetrix)Hold at 25 C
Quantity Used:
Amount15 microgram Fragmented cRNA
Time: 16 hours
Concentration: 50 pg/ml
Volume: 200 microlitres
Temperature: 45 C
Other Information:
Spike(Source: Affymetrix)Control Oligo B2 to 50pM for landing lights
Spike(Source: Affymetrix)20X Eukaryotic Hybridisation Controls (bioB (1.5pM, bioC 5pM, bioD 25pM, cre 100pM)
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

BioSource 1 Information

BioSource Name: Rayson_647-4_WT_6hr_cordycepin_Rep1_ATH1
Organism: Arabidopsis thaliana
Alias: Col-8 (Columbia)
Stock Code: N60000
Genetic Background: Col-0 (Columbia)
Age: 3 weeks
Growth Conditions:
StratificationImbibed in the dark at 4 °C for 3 days.
ProtocolPlants were grown in a Sanyo MLR-352 H with additive humidity under a typical short day cycle (8hr light:16 hr dark) The temperature was maintained at 21-23°C and the humidity was maintained at 65 % RH. Stratified seeds were sown onto SHL compost at a density of 4 seed per cell (24 cell tray). Trays were covered in a propagator lid, which was removed 7 days after sowing. Compost was kept moist throughout.
Soil ConstituentsSinclair Potting Growth Medium (www.william-sinclair.co.uk)
Plant Spacing1 per cell
waterCompost was kept moist at all times
Temperature22 °C average, 23 °C day, 21 °C night
Humidity65 % average, 65 % day, 65 % night
MediumNone
Lighting(Source: Cool white fluorescent manufactured by SANYO. Intensity: 175µEinsteins. Wavelength: White)
Developmental Stage:
Growth Stage(Source: Paradigm Genetics)
Genetic Variation: ecotype
Tissue: Aerial tissues
in vivo Treatment: Time = 6 hours Aerial tissues were harvested into incubation buffer (IB: 1 mM Pipes, 1 mM Sodium citrate, 1 mM Potassium chloride, 15 mM Sucrose). After 30 minutes in IB, all except the time (T) = 0 samples were transferred to IB with cordycepin (150 µg ml-1). The T = 0 samples were transferred to fresh IB without cordycepin. All flasks (including the T = 0) were transferred to a bell jar and vacuum infiltrated. Immediately post-infiltration was classed as T =0.
Additional Organism Information:
Sample DescriptionSoil-grown plant grown under short-day conditions.
Other Information:
Timecourse start procedureThe time-course started immediately after the vacuum infiltration.

Protocols for BioSource 1

Extraction Protocol Information

Name of Protocol: Qiagen / Trizol
Method:
ProtocolQiagen RNAeasy kit protocol
Type: total RNA
Amplification: Amplified during labelling process - see labelling protocol
Other Information:
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

Labelling Protocol Information

Name of Protocol: Biotin Labelled cRNA using Affymetrix Enzo Kit
Amount Labelled: Unknown
Label Name: Biotin
Labelling Method: In-vitro transcription method
Other Information:
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

Slide Information

Date Hybridised:

Array Design Information

Slide Design Name: Affymetrix ATH1 Arabidopsis Genome Array
Array Type: Affymetrix Gene Chip
Number of Spots on the Array: 22810

Scanning Protocol Information

Name: Affymetrix MAS 5.0 Standard Scanning
Width of Image Produced (Pixels): 4733
Height of Image Produced (Pixels): 4733
X Resolution (microns): 3
Y Resolution (microns): 3
Scanner Name: Agilent 2500A GeneArray Scanner
Software Name: Affymetrix Micro Array Suite 5.0

Image Analysis Information

Scaling Factor:1.315067
Name: Affymetrix MAS 5.0 Standard Image Analysis
Software Name: Affymetrix Microarray Analysis Suite 5.0

Normalisation Information

Strategy Name: Affymetrix MAS 5.0 Scaling Protocol
Algorithm: The top 2% and bottom 2% of signal intensities are excluded, then the mean is calculated. The original signal values are scaled such that the mean is made equal to 100.
Control Elements:
none
Normalisation Protocol Parameters:
Spike_AFFX-r2-P1-cre_3-5-ratio1.352094
Spike_AFFX-r2-Ec-bioB_3-5-ratio1.017426
Spike_AFFX-r2-Bs-dap_5_signal85.768814
NoiseAvg:2.75,Std:0.11,Min:2.4,Max:3.0
Control DirectionAntisense
Spike_AFFX-r2-Ec-bioB_5_detectionM
Spike_AFFX-r2-Ec-bioB_3_detectionP
Spike_AFFX-r2-Bs-thr_5_signal38.299809
#P10876
Spike_AFFX-r2-Bs-phe_M_signal55.743752
Corner-Avg:10173,Count:32
Spike_AFFX-r2-P1-cre_3_detectionP
Spike_AFFX-r2-Ec-bioB_M_signal23.902206
Spike_AFFX-r2-Ec-bioB_3_signal28.384745
Spike_AFFX-r2-Bs-lys_M_signal31.147072
Spike_AFFX-r2-P1-cre_3_signal10578.183594
Spike_AFFX-r2-Bs-lys_3-5-ratio1.156587
Spike_AFFX-r2-Bs-dap_M_signal291.132751
Spike_AFFX-r2-Bs-thr_M_detectionP
Probe Pair Threshold1
Spike_AFFX-r2-Bs-thr_3-5-ratio7.320441
Spike_AFFX-r2-Ec-bioB_avg-signal26.728510
Spike_AFFX-r2-Bs-thr_avg-signal157.181274
Spike_AFFX-r2-Bs-thr_5_detectionP
Spike_AFFX-r2-Bs-dap_3_detectionP
Spike_AFFX-r2-Ec-bioD_5_signal2065.182861
Spike_AFFX-r2-Bs-phe_5_signal48.598415
Spike_AFFX-r2-P1-cre_5_detectionP
Spike_AFFX-r2-Ec-bioD_3_signal2812.603516
RawQ2.759375
Spike_AFFX-r2-Bs-lys_5_signal46.053238
Signal(A)8.816825
%A49.614204
Signal(All)167.752548
Corner+Avg:171,Count:32
Spike_AFFX-r2-Ec-bioD_5_detectionP
Spike_AFFX-r2-Ec-bioC_3_signal695.194580
Spike_AFFX-r2-Bs-lys_3_detectionP
Spike_AFFX-r2-Bs-phe_3_signal82.237160
Spike_AFFX-r2-Ec-bioD_avg-signal2438.893066
Spike_AFFX-r2-Bs-dap_5_detectionP
Spike_AFFX-r2-Bs-phe_5_detectionP
%P47.680843
Spike_AFFX-r2-Bs-lys_avg-signal43.488300
Spike_AFFX-r2-P1-cre_avg-signal9200.871094
Spike_AFFX-r2-Ec-bioB_M_detectionP
Spike_AFFX-r2-Bs-lys_M_detectionP
#Probe Sets Exceeding Probe Pair Threshold22810
%M2.704954
Spike_AFFX-r2-Bs-thr_3_signal280.371490
Spike_AFFX-r2-Ec-bioC_5_detectionP
Spike_AFFX-r2-Bs-phe_3_detectionP
Spike_AFFX-r2-Ec-bioC_3_detectionP
Spike_AFFX-r2-Bs-phe_avg-signal62.193111
Spike_AFFX-r2-Bs-dap_3_signal541.476013
Spike_AFFX-r2-Ec-bioD_3-5-ratio1.361915
Spike_AFFX-r2-Ec-bioD_3_detectionP
Spike_AFFX-r2-Ec-bioC_5_signal660.803101
#M617
Spike_AFFX-r2-Bs-lys_5_detectionP
Spike_AFFX-r2-Bs-lys_3_signal53.264591
Spike_AFFX-r2-Bs-dap_M_detectionP
Spike_AFFX-r2-Ec-bioC_3-5-ratio1.052045
Spike_AFFX-r2-Bs-thr_M_signal152.872543
Signal(P)341.102997
Spike_AFFX-r2-Bs-phe_3-5-ratio1.692178
Spike_AFFX-r2-Bs-thr_3_detectionP
Central-Avg:9074,Count:9
Spike_AFFX-r2-P1-cre_5_signal7823.559082
Spike_AFFX-r2-Bs-phe_M_detectionP
#A11317
Signal(M)27.259844
BackgroundAvg:64.68,Std:0.45,Min:63.8,Max:66.5
Spike_AFFX-r2-Ec-bioC_avg-signal677.998840
Spike_AFFX-r2-Bs-dap_avg-signal306.125854
Spike_AFFX-r2-Bs-dap_3-5-ratio6.313204
Spike_AFFX-r2-Ec-bioB_5_signal27.898586
ScaleMaskAll
BG2
VZ4
TGT100
Alpha10.040000
Alpha20.060000
SF1.315067
NF1.000000
HZ4
Tau0.015000

Slide: Rayson_647-3_WT_3hr_cordycepin_Rep1_ATH1

Add to Slide Selection

Get the data!

Summary BioSource 1 Information Extraction / Labelling Protocol for Source 1 Information Slide Information Hybridisation Analysis Information for "Affymetrix MAS 5.0 Scaling Protocol" normalised data.

Summary Information

This slide has had 1 extract(s) hybridised to it (1-colour).

  • Extract 1 ("Rayson_647-3_WT_3hr_cordycepin_Rep1_ATH1") was made from "Arabidopsis thaliana" and labelled using "Biotin Labelled cRNA using Affymetrix Enzo Kit".

The slide used was "Affymetrix ATH1 Arabidopsis Genome Array".

Click on the options above for more information about this slide and its preparation.

Hybridisation Protocol Information

Name of Protocol: Affymetrix Antibody Amplification Protocol
Solution:
BufferEDTA, NaCl, Tween20 according to Affymetrix protocol
Water
Blocking Agent:
Herring Sperm DNA(Source: Promega)100pg/ml
Acetylated BSA(Source: Sigma-Aldrich)0.1mg/ml
Wash Procedure:
Step 1(Source: Affymetrix)10 cycles of 2 mixes/cycle with Non-Stringent buffer
Step 2(Source: Affymetrix)4 cycles of 15 mixes/cycle with Stringent buffer
Step 3(Source: Affymetrix)Stain the probe array for 10 minutes in Streptavaidin Phycoerythrin (SAPE) solution at 25 C
Step 4(Source: Affymetrix)10 cycles of 4 mixes/cycle with Non-Stringent buffer
Step 5(Source: Affymetrix)Stain the probe array for 10 minutes in antibody solution
Step 6(Source: Affymetrix)Stain the probe array for 10 minutes in SAPE solution at 25 C
Step 7(Source: Affymetrix)15 cycles of 4mixes/cycle with Non-Stringent buffer at 30 C
Step 8(Source: Affymetrix)Hold at 25 C
Quantity Used:
Amount15 microgram Fragmented cRNA
Time: 16 hours
Concentration: 50 pg/ml
Volume: 200 microlitres
Temperature: 45 C
Other Information:
Spike(Source: Affymetrix)Control Oligo B2 to 50pM for landing lights
Spike(Source: Affymetrix)20X Eukaryotic Hybridisation Controls (bioB (1.5pM, bioC 5pM, bioD 25pM, cre 100pM)
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

BioSource 1 Information

BioSource Name: Rayson_647-3_WT_3hr_cordycepin_Rep1_ATH1
Organism: Arabidopsis thaliana
Alias: Col-8 (Columbia)
Stock Code: N60000
Genetic Background: Col-0 (Columbia)
Age: 3 weeks
Growth Conditions:
StratificationImbibed in the dark at 4 °C for 3 days.
ProtocolPlants were grown in a Sanyo MLR-352 H with additive humidity under a typical short day cycle (8hr light:16 hr dark) The temperature was maintained at 21-23°C and the humidity was maintained at 65 % RH. Stratified seeds were sown onto SHL compost at a density of 4 seed per cell (24 cell tray). Trays were covered in a propagator lid, which was removed 7 days after sowing. Compost was kept moist throughout.
Soil ConstituentsSinclair Potting Growth Medium (www.william-sinclair.co.uk)
Plant Spacing1 per cell
waterCompost was kept moist at all times
Temperature22 °C average, 23 °C day, 21 °C night
Humidity65 % average, 65 % day, 65 % night
MediumNone
Lighting(Source: Cool white fluorescent manufactured by SANYO. Intensity: 175µEinsteins. Wavelength: White)
Developmental Stage:
Growth Stage(Source: Paradigm Genetics)
Genetic Variation: ecotype
Tissue: Aerial tissues
in vivo Treatment: Time = 3 hours Aerial tissues were harvested into incubation buffer (IB: 1 mM Pipes, 1 mM Sodium citrate, 1 mM Potassium chloride, 15 mM Sucrose). After 30 minutes in IB, all except the time (T) = 0 samples were transferred to IB with cordycepin (150 µg ml-1). The T = 0 samples were transferred to fresh IB without cordycepin. All flasks (including the T = 0) were transferred to a bell jar and vacuum infiltrated. Immediately post-infiltration was classed as T =0.
Additional Organism Information:
Sample DescriptionSoil-grown plant grown under short-day conditions.
Other Information:
Timecourse start procedureThe time-course started immediately after the vacuum infiltration.

Protocols for BioSource 1

Extraction Protocol Information

Name of Protocol: Qiagen / Trizol
Method:
ProtocolQiagen RNAeasy kit protocol
Type: total RNA
Amplification: Amplified during labelling process - see labelling protocol
Other Information:
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

Labelling Protocol Information

Name of Protocol: Biotin Labelled cRNA using Affymetrix Enzo Kit
Amount Labelled: Unknown
Label Name: Biotin
Labelling Method: In-vitro transcription method
Other Information:
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

Slide Information

Date Hybridised:

Array Design Information

Slide Design Name: Affymetrix ATH1 Arabidopsis Genome Array
Array Type: Affymetrix Gene Chip
Number of Spots on the Array: 22810

Scanning Protocol Information

Name: Affymetrix MAS 5.0 Standard Scanning
Width of Image Produced (Pixels): 4733
Height of Image Produced (Pixels): 4733
X Resolution (microns): 3
Y Resolution (microns): 3
Scanner Name: Agilent 2500A GeneArray Scanner
Software Name: Affymetrix Micro Array Suite 5.0

Image Analysis Information

Scaling Factor:0.79122
Name: Affymetrix MAS 5.0 Standard Image Analysis
Software Name: Affymetrix Microarray Analysis Suite 5.0

Normalisation Information

Strategy Name: Affymetrix MAS 5.0 Scaling Protocol
Algorithm: The top 2% and bottom 2% of signal intensities are excluded, then the mean is calculated. The original signal values are scaled such that the mean is made equal to 100.
Control Elements:
none
Normalisation Protocol Parameters:
Spike_AFFX-r2-P1-cre_3-5-ratio1.241364
Spike_AFFX-r2-Ec-bioB_3-5-ratio1.367012
Spike_AFFX-r2-Bs-dap_5_signal37.914421
NoiseAvg:3.12,Std:0.14,Min:2.7,Max:3.5
Control DirectionAntisense
Spike_AFFX-r2-Ec-bioB_5_detectionP
Spike_AFFX-r2-Ec-bioB_3_detectionP
Spike_AFFX-r2-Bs-thr_5_signal21.069260
#P11992
Spike_AFFX-r2-Bs-phe_M_signal36.031570
Corner-Avg:11559,Count:32
Spike_AFFX-r2-P1-cre_3_detectionP
Spike_AFFX-r2-Ec-bioB_M_signal18.351500
Spike_AFFX-r2-Ec-bioB_3_signal19.112324
Spike_AFFX-r2-Bs-lys_M_signal17.091881
Spike_AFFX-r2-P1-cre_3_signal7000.560059
Spike_AFFX-r2-Bs-lys_3-5-ratio1.562043
Spike_AFFX-r2-Bs-dap_M_signal192.144379
Spike_AFFX-r2-Bs-thr_M_detectionP
Probe Pair Threshold1
Spike_AFFX-r2-Bs-thr_3-5-ratio7.672863
Spike_AFFX-r2-Ec-bioB_avg-signal17.148306
Spike_AFFX-r2-Bs-thr_avg-signal81.216080
Spike_AFFX-r2-Bs-thr_5_detectionP
Spike_AFFX-r2-Bs-dap_3_detectionP
Spike_AFFX-r2-Ec-bioD_5_signal1708.853149
Spike_AFFX-r2-Bs-phe_5_signal18.684374
Spike_AFFX-r2-P1-cre_5_detectionP
Spike_AFFX-r2-Ec-bioD_3_signal2290.745117
RawQ2.840178
Spike_AFFX-r2-Bs-lys_5_signal20.551277
Signal(A)6.185685
%A45.221394
Signal(All)158.876587
Corner+Avg:213,Count:32
Spike_AFFX-r2-Ec-bioD_5_detectionP
Spike_AFFX-r2-Ec-bioC_3_signal454.371796
Spike_AFFX-r2-Bs-lys_3_detectionP
Spike_AFFX-r2-Bs-phe_3_signal62.314526
Spike_AFFX-r2-Ec-bioD_avg-signal1999.799072
Spike_AFFX-r2-Bs-dap_5_detectionP
Spike_AFFX-r2-Bs-phe_5_detectionP
%P52.573433
Spike_AFFX-r2-Bs-lys_avg-signal23.248377
Spike_AFFX-r2-P1-cre_avg-signal6319.985352
Spike_AFFX-r2-Ec-bioB_M_detectionP
Spike_AFFX-r2-Bs-lys_M_detectionP
#Probe Sets Exceeding Probe Pair Threshold22810
%M2.205173
Spike_AFFX-r2-Bs-thr_3_signal161.661545
Spike_AFFX-r2-Ec-bioC_5_detectionP
Spike_AFFX-r2-Bs-phe_3_detectionP
Spike_AFFX-r2-Ec-bioC_3_detectionP
Spike_AFFX-r2-Bs-phe_avg-signal39.010159
Spike_AFFX-r2-Bs-dap_3_signal330.649078
Spike_AFFX-r2-Ec-bioD_3-5-ratio1.340516
Spike_AFFX-r2-Ec-bioD_3_detectionP
Spike_AFFX-r2-Ec-bioC_5_signal430.110046
#M503
Spike_AFFX-r2-Bs-lys_5_detectionP
Spike_AFFX-r2-Bs-lys_3_signal32.101974
Spike_AFFX-r2-Bs-dap_M_detectionP
Spike_AFFX-r2-Ec-bioC_3-5-ratio1.056408
Spike_AFFX-r2-Bs-thr_M_signal60.917431
Signal(P)296.000305
Spike_AFFX-r2-Bs-phe_3-5-ratio3.335114
Spike_AFFX-r2-Bs-thr_3_detectionP
Central-Avg:10159,Count:9
Spike_AFFX-r2-P1-cre_5_signal5639.410645
Spike_AFFX-r2-Bs-phe_M_detectionP
#A10315
Signal(M)20.942408
BackgroundAvg:70.34,Std:0.67,Min:68.8,Max:73.0
Spike_AFFX-r2-Ec-bioC_avg-signal442.240906
Spike_AFFX-r2-Bs-dap_avg-signal186.902634
Spike_AFFX-r2-Bs-dap_3-5-ratio8.720932
Spike_AFFX-r2-Ec-bioB_5_signal13.981094
ScaleMaskAll
BG2
VZ4
TGT100
Alpha10.040000
Alpha20.060000
SF0.791220
NF1.000000
HZ4
Tau0.015000

Slide: Rayson_647-2_WT_1hr_cordycepin_Rep1_ATH1

Add to Slide Selection

Get the data!

Summary BioSource 1 Information Extraction / Labelling Protocol for Source 1 Information Slide Information Hybridisation Analysis Information for "Affymetrix MAS 5.0 Scaling Protocol" normalised data.

Summary Information

This slide has had 1 extract(s) hybridised to it (1-colour).

  • Extract 1 ("Rayson_647-2_WT_1hr_cordycepin_Rep1_ATH1") was made from "Arabidopsis thaliana" and labelled using "Biotin Labelled cRNA using Affymetrix Enzo Kit".

The slide used was "Affymetrix ATH1 Arabidopsis Genome Array".

Click on the options above for more information about this slide and its preparation.

Hybridisation Protocol Information

Name of Protocol: Affymetrix Antibody Amplification Protocol
Solution:
BufferEDTA, NaCl, Tween20 according to Affymetrix protocol
Water
Blocking Agent:
Herring Sperm DNA(Source: Promega)100pg/ml
Acetylated BSA(Source: Sigma-Aldrich)0.1mg/ml
Wash Procedure:
Step 1(Source: Affymetrix)10 cycles of 2 mixes/cycle with Non-Stringent buffer
Step 2(Source: Affymetrix)4 cycles of 15 mixes/cycle with Stringent buffer
Step 3(Source: Affymetrix)Stain the probe array for 10 minutes in Streptavaidin Phycoerythrin (SAPE) solution at 25 C
Step 4(Source: Affymetrix)10 cycles of 4 mixes/cycle with Non-Stringent buffer
Step 5(Source: Affymetrix)Stain the probe array for 10 minutes in antibody solution
Step 6(Source: Affymetrix)Stain the probe array for 10 minutes in SAPE solution at 25 C
Step 7(Source: Affymetrix)15 cycles of 4mixes/cycle with Non-Stringent buffer at 30 C
Step 8(Source: Affymetrix)Hold at 25 C
Quantity Used:
Amount15 microgram Fragmented cRNA
Time: 16 hours
Concentration: 50 pg/ml
Volume: 200 microlitres
Temperature: 45 C
Other Information:
Spike(Source: Affymetrix)Control Oligo B2 to 50pM for landing lights
Spike(Source: Affymetrix)20X Eukaryotic Hybridisation Controls (bioB (1.5pM, bioC 5pM, bioD 25pM, cre 100pM)
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

BioSource 1 Information

BioSource Name: Rayson_647-2_WT_1hr_cordycepin_Rep1_ATH1
Organism: Arabidopsis thaliana
Alias: Col-8 (Columbia)
Stock Code: N60000
Genetic Background: Col-0 (Columbia)
Age: 3 weeks
Growth Conditions:
StratificationImbibed in the dark at 4 °C for 3 days.
ProtocolPlants were grown in a Sanyo MLR-352 H with additive humidity under a typical short day cycle (8hr light:16 hr dark) The temperature was maintained at 21-23°C and the humidity was maintained at 65 % RH. Stratified seeds were sown onto SHL compost at a density of 4 seed per cell (24 cell tray). Trays were covered in a propagator lid, which was removed 7 days after sowing. Compost was kept moist throughout.
Soil ConstituentsSinclair Potting Growth Medium (www.william-sinclair.co.uk)
Plant Spacing1 per cell
waterCompost was kept moist at all times
Temperature22 °C average, 23 °C day, 21 °C night
Humidity65 % average, 65 % day, 65 % night
MediumNone
Lighting(Source: Cool white fluorescent manufactured by SANYO. Intensity: 175µEinsteins. Wavelength: White)
Developmental Stage:
Growth Stage(Source: Paradigm Genetics)
Genetic Variation: ecotype
Tissue: Aerial tissues
in vivo Treatment: Time = 1 hour Aerial tissues were harvested into incubation buffer (IB: 1 mM Pipes, 1 mM Sodium citrate, 1 mM Potassium chloride, 15 mM Sucrose). After 30 minutes in IB, all except the time (T) = 0 samples were transferred to IB with cordycepin (150 µg ml-1). The T = 0 samples were transferred to fresh IB without cordycepin. All flasks (including the T = 0) were transferred to a bell jar and vacuum infiltrated. Immediately post-infiltration was classed as T =0.
Additional Organism Information:
Sample DescriptionSoil-grown plant grown under short-day conditions.
Other Information:
Timecourse start procedureThe time-course started immediately after the vacuum infiltration.

Protocols for BioSource 1

Extraction Protocol Information

Name of Protocol: Qiagen / Trizol
Method:
ProtocolQiagen RNAeasy kit protocol
Type: total RNA
Amplification: Amplified during labelling process - see labelling protocol
Other Information:
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

Labelling Protocol Information

Name of Protocol: Biotin Labelled cRNA using Affymetrix Enzo Kit
Amount Labelled: Unknown
Label Name: Biotin
Labelling Method: In-vitro transcription method
Other Information:
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

Slide Information

Date Hybridised:

Array Design Information

Slide Design Name: Affymetrix ATH1 Arabidopsis Genome Array
Array Type: Affymetrix Gene Chip
Number of Spots on the Array: 22810

Scanning Protocol Information

Name: Affymetrix MAS 5.0 Standard Scanning
Width of Image Produced (Pixels): 4733
Height of Image Produced (Pixels): 4733
X Resolution (microns): 3
Y Resolution (microns): 3
Scanner Name: Agilent 2500A GeneArray Scanner
Software Name: Affymetrix Micro Array Suite 5.0

Image Analysis Information

Scaling Factor:0.56567
Name: Affymetrix MAS 5.0 Standard Image Analysis
Software Name: Affymetrix Microarray Analysis Suite 5.0

Normalisation Information

Strategy Name: Affymetrix MAS 5.0 Scaling Protocol
Algorithm: The top 2% and bottom 2% of signal intensities are excluded, then the mean is calculated. The original signal values are scaled such that the mean is made equal to 100.
Control Elements:
none
Normalisation Protocol Parameters:
Spike_AFFX-r2-P1-cre_3-5-ratio1.423198
Spike_AFFX-r2-Ec-bioB_3-5-ratio1.212877
Spike_AFFX-r2-Bs-dap_5_signal32.355068
NoiseAvg:3.38,Std:0.11,Min:3.1,Max:3.7
Control DirectionAntisense
Spike_AFFX-r2-Ec-bioB_5_detectionP
Spike_AFFX-r2-Ec-bioB_3_detectionP
Spike_AFFX-r2-Bs-thr_5_signal14.614036
#P12848
Spike_AFFX-r2-Bs-phe_M_signal34.412624
Corner-Avg:13230,Count:32
Spike_AFFX-r2-P1-cre_3_detectionP
Spike_AFFX-r2-Ec-bioB_M_signal12.025096
Spike_AFFX-r2-Ec-bioB_3_signal8.568262
Spike_AFFX-r2-Bs-lys_M_signal20.030087
Spike_AFFX-r2-P1-cre_3_signal5411.574219
Spike_AFFX-r2-Bs-lys_3-5-ratio1.958049
Spike_AFFX-r2-Bs-dap_M_signal173.702362
Spike_AFFX-r2-Bs-thr_M_detectionP
Probe Pair Threshold1
Spike_AFFX-r2-Bs-thr_3-5-ratio5.788400
Spike_AFFX-r2-Ec-bioB_avg-signal9.219256
Spike_AFFX-r2-Bs-thr_avg-signal47.483139
Spike_AFFX-r2-Bs-thr_5_detectionP
Spike_AFFX-r2-Bs-dap_3_detectionP
Spike_AFFX-r2-Ec-bioD_5_signal1311.510620
Spike_AFFX-r2-Bs-phe_5_signal20.734392
Spike_AFFX-r2-P1-cre_5_detectionP
Spike_AFFX-r2-Ec-bioD_3_signal1897.286621
RawQ3.003758
Spike_AFFX-r2-Bs-lys_5_signal25.113989
Signal(A)4.628379
%A41.836914
Signal(All)150.226486
Corner+Avg:230,Count:32
Spike_AFFX-r2-Ec-bioD_5_detectionP
Spike_AFFX-r2-Ec-bioC_3_signal354.350037
Spike_AFFX-r2-Bs-lys_3_detectionP
Spike_AFFX-r2-Bs-phe_3_signal53.673965
Spike_AFFX-r2-Ec-bioD_avg-signal1604.398682
Spike_AFFX-r2-Bs-dap_5_detectionP
Spike_AFFX-r2-Bs-phe_5_detectionP
%P56.326172
Spike_AFFX-r2-Bs-lys_avg-signal31.439497
Spike_AFFX-r2-P1-cre_avg-signal4606.989258
Spike_AFFX-r2-Ec-bioB_M_detectionP
Spike_AFFX-r2-Bs-lys_M_detectionP
#Probe Sets Exceeding Probe Pair Threshold22810
%M1.836914
Spike_AFFX-r2-Bs-thr_3_signal84.591881
Spike_AFFX-r2-Ec-bioC_5_detectionP
Spike_AFFX-r2-Bs-phe_3_detectionP
Spike_AFFX-r2-Ec-bioC_3_detectionP
Spike_AFFX-r2-Bs-phe_avg-signal36.273663
Spike_AFFX-r2-Bs-dap_3_signal285.765259
Spike_AFFX-r2-Ec-bioD_3-5-ratio1.446642
Spike_AFFX-r2-Ec-bioD_3_detectionP
Spike_AFFX-r2-Ec-bioC_5_signal334.369812
#M419
Spike_AFFX-r2-Bs-lys_5_detectionP
Spike_AFFX-r2-Bs-lys_3_signal49.174412
Spike_AFFX-r2-Bs-dap_M_detectionP
Spike_AFFX-r2-Ec-bioC_3-5-ratio1.059755
Spike_AFFX-r2-Bs-thr_M_signal43.243507
Signal(P)262.724548
Spike_AFFX-r2-Bs-phe_3-5-ratio2.588644
Spike_AFFX-r2-Bs-thr_3_detectionP
Central-Avg:11219,Count:9
Spike_AFFX-r2-P1-cre_5_signal3802.404785
Spike_AFFX-r2-Bs-phe_M_detectionP
#A9543
Signal(M)16.736225
BackgroundAvg:74.70,Std:0.36,Min:74.0,Max:76.3
Spike_AFFX-r2-Ec-bioC_avg-signal344.359924
Spike_AFFX-r2-Bs-dap_avg-signal163.940903
Spike_AFFX-r2-Bs-dap_3-5-ratio8.832164
Spike_AFFX-r2-Ec-bioB_5_signal7.064414
ScaleMaskAll
BG2
VZ4
TGT100
Alpha10.040000
Alpha20.060000
SF0.565670
NF1.000000
HZ4
Tau0.015000

Slide: Rayson_647-1_WT_0hr_cordycepin_Rep1_ATH1

Add to Slide Selection

Get the data!

Summary BioSource 1 Information Extraction / Labelling Protocol for Source 1 Information Slide Information Hybridisation Analysis Information for "Affymetrix MAS 5.0 Scaling Protocol" normalised data.

Summary Information

This slide has had 1 extract(s) hybridised to it (1-colour).

  • Extract 1 ("Rayson_647-1_WT_0hr_cordycepin_Rep1_ATH1") was made from "Arabidopsis thaliana" and labelled using "Biotin Labelled cRNA using Affymetrix Enzo Kit".

The slide used was "Affymetrix ATH1 Arabidopsis Genome Array".

Click on the options above for more information about this slide and its preparation.

Hybridisation Protocol Information

Name of Protocol: Affymetrix Antibody Amplification Protocol
Solution:
BufferEDTA, NaCl, Tween20 according to Affymetrix protocol
Water
Blocking Agent:
Herring Sperm DNA(Source: Promega)100pg/ml
Acetylated BSA(Source: Sigma-Aldrich)0.1mg/ml
Wash Procedure:
Step 1(Source: Affymetrix)10 cycles of 2 mixes/cycle with Non-Stringent buffer
Step 2(Source: Affymetrix)4 cycles of 15 mixes/cycle with Stringent buffer
Step 3(Source: Affymetrix)Stain the probe array for 10 minutes in Streptavaidin Phycoerythrin (SAPE) solution at 25 C
Step 4(Source: Affymetrix)10 cycles of 4 mixes/cycle with Non-Stringent buffer
Step 5(Source: Affymetrix)Stain the probe array for 10 minutes in antibody solution
Step 6(Source: Affymetrix)Stain the probe array for 10 minutes in SAPE solution at 25 C
Step 7(Source: Affymetrix)15 cycles of 4mixes/cycle with Non-Stringent buffer at 30 C
Step 8(Source: Affymetrix)Hold at 25 C
Quantity Used:
Amount15 microgram Fragmented cRNA
Time: 16 hours
Concentration: 50 pg/ml
Volume: 200 microlitres
Temperature: 45 C
Other Information:
Spike(Source: Affymetrix)Control Oligo B2 to 50pM for landing lights
Spike(Source: Affymetrix)20X Eukaryotic Hybridisation Controls (bioB (1.5pM, bioC 5pM, bioD 25pM, cre 100pM)
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

BioSource 1 Information

BioSource Name: Rayson_647-1_WT_0hr_cordycepin_Rep1_ATH1
Organism: Arabidopsis thaliana
Alias: Col-8 (Columbia)
Stock Code: N60000
Genetic Background: Col-0 (Columbia)
Age: 3 weeks
Growth Conditions:
StratificationImbibed in the dark at 4 °C for 3 days.
ProtocolPlants were grown in a Sanyo MLR-352 H with additive humidity under a typical short day cycle (8hr light:16 hr dark) The temperature was maintained at 21-23°C and the humidity was maintained at 65 % RH. Stratified seeds were sown onto SHL compost at a density of 4 seed per cell (24 cell tray). Trays were covered in a propagator lid, which was removed 7 days after sowing. Compost was kept moist throughout.
Soil ConstituentsSinclair Potting Growth Medium (www.william-sinclair.co.uk)
Plant Spacing1 per cell
waterCompost was kept moist at all times
Temperature22 °C average, 23 °C day, 21 °C night
Humidity65 % average, 65 % day, 65 % night
MediumNone
Lighting(Source: Cool white fluorescent manufactured by SANYO. Intensity: 175µEinsteins. Wavelength: White)
Developmental Stage:
Growth Stage(Source: Paradigm Genetics)
Genetic Variation: ecotype
Tissue: Aerial tissues
in vivo Treatment: Time = 0 (no cordycepin) Aerial tissues were harvested into incubation buffer (IB: 1 mM Pipes, 1 mM Sodium citrate, 1 mM Potassium chloride, 15 mM Sucrose). After 30 minutes in IB, all except the time (T) = 0 samples were transferred to IB with cordycepin (150 µg ml-1). The T = 0 samples were transferred to fresh IB without cordycepin. All flasks (including the T = 0) were transferred to a bell jar and vacuum infiltrated. Immediately post-infiltration was classed as T =0.
Additional Organism Information:
Sample DescriptionSoil-grown plant grown under short-day conditions.
Other Information:
Timecourse start procedureThe time-course started immediately after the vacuum infiltration.

Protocols for BioSource 1

Extraction Protocol Information

Name of Protocol: Qiagen / Trizol
Method:
ProtocolQiagen RNAeasy kit protocol
Type: total RNA
Amplification: Amplified during labelling process - see labelling protocol
Other Information:
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

Labelling Protocol Information

Name of Protocol: Biotin Labelled cRNA using Affymetrix Enzo Kit
Amount Labelled: Unknown
Label Name: Biotin
Labelling Method: In-vitro transcription method
Other Information:
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

Slide Information

Date Hybridised:

Array Design Information

Slide Design Name: Affymetrix ATH1 Arabidopsis Genome Array
Array Type: Affymetrix Gene Chip
Number of Spots on the Array: 22810

Scanning Protocol Information

Name: Affymetrix MAS 5.0 Standard Scanning
Width of Image Produced (Pixels): 4733
Height of Image Produced (Pixels): 4733
X Resolution (microns): 3
Y Resolution (microns): 3
Scanner Name: Agilent 2500A GeneArray Scanner
Software Name: Affymetrix Micro Array Suite 5.0

Image Analysis Information

Scaling Factor:0.517827
Name: Affymetrix MAS 5.0 Standard Image Analysis
Software Name: Affymetrix Microarray Analysis Suite 5.0

Normalisation Information

Strategy Name: Affymetrix MAS 5.0 Scaling Protocol
Algorithm: The top 2% and bottom 2% of signal intensities are excluded, then the mean is calculated. The original signal values are scaled such that the mean is made equal to 100.
Control Elements:
none
Normalisation Protocol Parameters:
Spike_AFFX-r2-P1-cre_3-5-ratio1.367543
Spike_AFFX-r2-Ec-bioB_3-5-ratio0.507777
Spike_AFFX-r2-Bs-dap_5_signal16.058538
NoiseAvg:3.61,Std:0.18,Min:3.2,Max:4.1
Control DirectionAntisense
Spike_AFFX-r2-Ec-bioB_5_detectionP
Spike_AFFX-r2-Ec-bioB_3_detectionP
Spike_AFFX-r2-Bs-thr_5_signal14.776528
#P13779
Spike_AFFX-r2-Bs-phe_M_signal17.786697
Corner-Avg:12445,Count:32
Spike_AFFX-r2-P1-cre_3_detectionP
Spike_AFFX-r2-Ec-bioB_M_signal14.913528
Spike_AFFX-r2-Ec-bioB_3_signal12.496088
Spike_AFFX-r2-Bs-lys_M_signal12.133327
Spike_AFFX-r2-P1-cre_3_signal5529.157227
Spike_AFFX-r2-Bs-lys_3-5-ratio2.183164
Spike_AFFX-r2-Bs-dap_M_signal95.071350
Spike_AFFX-r2-Bs-thr_M_detectionP
Probe Pair Threshold1
Spike_AFFX-r2-Bs-thr_3-5-ratio5.154696
Spike_AFFX-r2-Ec-bioB_avg-signal17.339674
Spike_AFFX-r2-Bs-thr_avg-signal41.479031
Spike_AFFX-r2-Bs-thr_5_detectionM
Spike_AFFX-r2-Bs-dap_3_detectionP
Spike_AFFX-r2-Ec-bioD_5_signal1209.095459
Spike_AFFX-r2-Bs-phe_5_signal14.589298
Spike_AFFX-r2-P1-cre_5_detectionP
Spike_AFFX-r2-Ec-bioD_3_signal1732.761597
RawQ3.050946
Spike_AFFX-r2-Bs-lys_5_signal13.350626
Signal(A)4.462904
%A37.702763
Signal(All)144.396027
Corner+Avg:221,Count:32
Spike_AFFX-r2-Ec-bioD_5_detectionP
Spike_AFFX-r2-Ec-bioC_3_signal346.258545
Spike_AFFX-r2-Bs-lys_3_detectionP
Spike_AFFX-r2-Bs-phe_3_signal41.018166
Spike_AFFX-r2-Ec-bioD_avg-signal1470.928467
Spike_AFFX-r2-Bs-dap_5_detectionP
Spike_AFFX-r2-Bs-phe_5_detectionP
%P60.407715
Spike_AFFX-r2-Bs-lys_avg-signal18.210188
Spike_AFFX-r2-P1-cre_avg-signal4786.145508
Spike_AFFX-r2-Ec-bioB_M_detectionP
Spike_AFFX-r2-Bs-lys_M_detectionP
#Probe Sets Exceeding Probe Pair Threshold22810
%M1.889522
Spike_AFFX-r2-Bs-thr_3_signal76.168503
Spike_AFFX-r2-Ec-bioC_5_detectionP
Spike_AFFX-r2-Bs-phe_3_detectionP
Spike_AFFX-r2-Ec-bioC_3_detectionP
Spike_AFFX-r2-Bs-phe_avg-signal24.464722
Spike_AFFX-r2-Bs-dap_3_signal212.450851
Spike_AFFX-r2-Ec-bioD_3-5-ratio1.433106
Spike_AFFX-r2-Ec-bioD_3_detectionP
Spike_AFFX-r2-Ec-bioC_5_signal323.567657
#M431
Spike_AFFX-r2-Bs-lys_5_detectionP
Spike_AFFX-r2-Bs-lys_3_signal29.146606
Spike_AFFX-r2-Bs-dap_M_detectionP
Spike_AFFX-r2-Ec-bioC_3-5-ratio1.070127
Spike_AFFX-r2-Bs-thr_M_signal33.492065
Signal(P)235.734299
Spike_AFFX-r2-Bs-phe_3-5-ratio2.811524
Spike_AFFX-r2-Bs-thr_3_detectionP
Central-Avg:10686,Count:9
Spike_AFFX-r2-P1-cre_5_signal4043.133545
Spike_AFFX-r2-Bs-phe_M_detectionP
#A8600
Signal(M)16.494999
BackgroundAvg:74.43,Std:0.65,Min:73.1,Max:76.4
Spike_AFFX-r2-Ec-bioC_avg-signal334.913086
Spike_AFFX-r2-Bs-dap_avg-signal107.860252
Spike_AFFX-r2-Bs-dap_3-5-ratio13.229775
Spike_AFFX-r2-Ec-bioB_5_signal24.609404
ScaleMaskAll
BG2
VZ4
TGT100
Alpha10.040000
Alpha20.060000
SF0.517827
NF1.000000
HZ4
Tau0.015000

Slide: Rayson_647-20_upf3-1_NMD_6hr _cordycepin_Rep1_ATH1

Add to Slide Selection

Get the data!

Summary BioSource 1 Information Extraction / Labelling Protocol for Source 1 Information Slide Information Hybridisation Analysis Information for "Affymetrix MAS 5.0 Scaling Protocol" normalised data.

Summary Information

This slide has had 1 extract(s) hybridised to it (1-colour).

  • Extract 1 ("Rayson_647-20_upf3-1_NMD_6hr _cordycepin_Rep1_ATH1") was made from "Arabidopsis thaliana" and labelled using "Biotin Labelled cRNA using Affymetrix Enzo Kit".

The slide used was "Affymetrix ATH1 Arabidopsis Genome Array".

Click on the options above for more information about this slide and its preparation.

Hybridisation Protocol Information

Name of Protocol: Affymetrix Antibody Amplification Protocol
Solution:
BufferEDTA, NaCl, Tween20 according to Affymetrix protocol
Water
Blocking Agent:
Herring Sperm DNA(Source: Promega)100pg/ml
Acetylated BSA(Source: Sigma-Aldrich)0.1mg/ml
Wash Procedure:
Step 1(Source: Affymetrix)10 cycles of 2 mixes/cycle with Non-Stringent buffer
Step 2(Source: Affymetrix)4 cycles of 15 mixes/cycle with Stringent buffer
Step 3(Source: Affymetrix)Stain the probe array for 10 minutes in Streptavaidin Phycoerythrin (SAPE) solution at 25 C
Step 4(Source: Affymetrix)10 cycles of 4 mixes/cycle with Non-Stringent buffer
Step 5(Source: Affymetrix)Stain the probe array for 10 minutes in antibody solution
Step 6(Source: Affymetrix)Stain the probe array for 10 minutes in SAPE solution at 25 C
Step 7(Source: Affymetrix)15 cycles of 4mixes/cycle with Non-Stringent buffer at 30 C
Step 8(Source: Affymetrix)Hold at 25 C
Quantity Used:
Amount15 microgram Fragmented cRNA
Time: 16 hours
Concentration: 50 pg/ml
Volume: 200 microlitres
Temperature: 45 C
Other Information:
Spike(Source: Affymetrix)Control Oligo B2 to 50pM for landing lights
Spike(Source: Affymetrix)20X Eukaryotic Hybridisation Controls (bioB (1.5pM, bioC 5pM, bioD 25pM, cre 100pM)
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

BioSource 1 Information

BioSource Name: Rayson_647-20_upf3-1_NMD_6hr _cordycepin_Rep1_ATH1
Organism: Arabidopsis thaliana
Alias: upf3-1
Stock Code: N9900
Genetic Background: Col-0
Age: 3 weeks
Growth Conditions:
StratificationImbibed in the dark at 4 °C for 3 days.
ProtocolPlants were grown in a Sanyo MLR-352 H with additive humidity under a typical short day cycle (8hr light:16 hr dark) The temperature was maintained at 21-23°C and the humidity was maintained at 65 % RH. Stratified seeds were sown onto SHL compost at a density of 4 seed per cell (24 cell tray). Trays were covered in a propagator lid, which was removed 7 days after sowing. Compost was kept moist throughout.
Soil ConstituentsSinclair Potting Growth Medium (www.william-sinclair.co.uk)
Plant Spacing1 per cell
waterCompost was kept moist at all times
Temperature22 °C average, 23 °C day, 21 °C night
Humidity65 % average, 65 % day, 65 % night
MediumNone
Lighting(Source: Cool white fluorescent manufactured by SANYO. Intensity: 175µEinsteins. Wavelength: White)
Developmental Stage:
Growth Stage(Source: Paradigm Genetics)
Tissue: Aerial tissues
in vivo Treatment: Time = 6 hours Aerial tissues were harvested into incubation buffer (IB: 1 mM Pipes, 1 mM Sodium citrate, 1 mM Potassium chloride, 15 mM Sucrose). After 30 minutes in IB, all except the time (T) = 0 samples were transferred to IB with cordycepin (150 µg ml-1). The T = 0 samples were transferred to fresh IB without cordycepin. All flasks (including the T = 0) were transferred to a bell jar and vacuum infiltrated. Immediately post-infiltration was classed as T =0.
Additional Organism Information:
Sample DescriptionSoil-grown plant grown under short-day conditions.
Other Information:
Timecourse start procedureThe time-course started immediately after the vacuum infiltration.

Protocols for BioSource 1

Extraction Protocol Information

Name of Protocol: Qiagen / Trizol
Method:
ProtocolQiagen RNAeasy kit protocol
Type: total RNA
Amplification: Amplified during labelling process - see labelling protocol
Other Information:
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

Labelling Protocol Information

Name of Protocol: Biotin Labelled cRNA using Affymetrix Enzo Kit
Amount Labelled: Unknown
Label Name: Biotin
Labelling Method: In-vitro transcription method
Other Information:
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

Slide Information

Date Hybridised:

Array Design Information

Slide Design Name: Affymetrix ATH1 Arabidopsis Genome Array
Array Type: Affymetrix Gene Chip
Number of Spots on the Array: 22810

Scanning Protocol Information

Name: Affymetrix MAS 5.0 Standard Scanning
Width of Image Produced (Pixels): 4733
Height of Image Produced (Pixels): 4733
X Resolution (microns): 3
Y Resolution (microns): 3
Scanner Name: Agilent 2500A GeneArray Scanner
Software Name: Affymetrix Micro Array Suite 5.0

Image Analysis Information

Scaling Factor:0.970609
Name: Affymetrix MAS 5.0 Standard Image Analysis
Software Name: Affymetrix Microarray Analysis Suite 5.0

Normalisation Information

Strategy Name: Affymetrix MAS 5.0 Scaling Protocol
Algorithm: The top 2% and bottom 2% of signal intensities are excluded, then the mean is calculated. The original signal values are scaled such that the mean is made equal to 100.
Control Elements:
none
Normalisation Protocol Parameters:
Spike_AFFX-r2-P1-cre_3-5-ratio1.481525
Spike_AFFX-r2-Ec-bioB_3-5-ratio1.268127
Spike_AFFX-r2-Bs-dap_5_signal67.988670
NoiseAvg:2.63,Std:0.06,Min:2.4,Max:2.8
Control DirectionAntisense
Spike_AFFX-r2-Ec-bioB_5_detectionP
Spike_AFFX-r2-Ec-bioB_3_detectionM
Spike_AFFX-r2-Bs-thr_5_signal31.402487
#P12454
Spike_AFFX-r2-Bs-phe_M_signal60.288330
Corner-Avg:11719,Count:32
Spike_AFFX-r2-P1-cre_3_detectionP
Spike_AFFX-r2-Ec-bioB_M_signal16.502480
Spike_AFFX-r2-Ec-bioB_3_signal28.500216
Spike_AFFX-r2-Bs-lys_M_signal35.811287
Spike_AFFX-r2-P1-cre_3_signal7185.640625
Spike_AFFX-r2-Bs-lys_3-5-ratio1.274238
Spike_AFFX-r2-Bs-dap_M_signal301.353088
Spike_AFFX-r2-Bs-thr_M_detectionP
Probe Pair Threshold1
Spike_AFFX-r2-Bs-thr_3-5-ratio9.320683
Spike_AFFX-r2-Ec-bioB_avg-signal22.492319
Spike_AFFX-r2-Bs-thr_avg-signal147.270935
Spike_AFFX-r2-Bs-thr_5_detectionP
Spike_AFFX-r2-Bs-dap_3_detectionP
Spike_AFFX-r2-Ec-bioD_5_signal1514.133911
Spike_AFFX-r2-Bs-phe_5_signal48.391315
Spike_AFFX-r2-P1-cre_5_detectionP
Spike_AFFX-r2-Ec-bioD_3_signal2179.120605
RawQ2.503789
Spike_AFFX-r2-Bs-lys_5_signal48.570480
Signal(A)6.204193
%A43.134590
Signal(All)154.500504
Corner+Avg:197,Count:32
Spike_AFFX-r2-Ec-bioD_5_detectionP
Spike_AFFX-r2-Ec-bioC_3_signal453.327820
Spike_AFFX-r2-Bs-lys_3_detectionP
Spike_AFFX-r2-Bs-phe_3_signal99.137146
Spike_AFFX-r2-Ec-bioD_avg-signal1846.627197
Spike_AFFX-r2-Bs-dap_5_detectionP
Spike_AFFX-r2-Bs-phe_5_detectionP
%P54.598862
Spike_AFFX-r2-Bs-lys_avg-signal48.757370
Spike_AFFX-r2-P1-cre_avg-signal6017.902344
Spike_AFFX-r2-Ec-bioB_M_detectionP
Spike_AFFX-r2-Bs-lys_M_detectionP
#Probe Sets Exceeding Probe Pair Threshold22810
%M2.266550
Spike_AFFX-r2-Bs-thr_3_signal292.692596
Spike_AFFX-r2-Ec-bioC_5_detectionP
Spike_AFFX-r2-Bs-phe_3_detectionP
Spike_AFFX-r2-Ec-bioC_3_detectionP
Spike_AFFX-r2-Bs-phe_avg-signal69.272263
Spike_AFFX-r2-Bs-dap_3_signal615.251038
Spike_AFFX-r2-Ec-bioD_3-5-ratio1.439186
Spike_AFFX-r2-Ec-bioD_3_detectionP
Spike_AFFX-r2-Ec-bioC_5_signal469.376801
#M517
Spike_AFFX-r2-Bs-lys_5_detectionP
Spike_AFFX-r2-Bs-lys_3_signal61.890347
Spike_AFFX-r2-Bs-dap_M_detectionP
Spike_AFFX-r2-Ec-bioC_3-5-ratio0.965808
Spike_AFFX-r2-Bs-thr_M_signal117.717712
Signal(P)277.215912
Spike_AFFX-r2-Bs-phe_3-5-ratio2.048656
Spike_AFFX-r2-Bs-thr_3_detectionP
Central-Avg:9867,Count:9
Spike_AFFX-r2-P1-cre_5_signal4850.163574
Spike_AFFX-r2-Bs-phe_M_detectionP
#A9839
Signal(M)20.631569
BackgroundAvg:60.83,Std:0.43,Min:60.0,Max:62.0
Spike_AFFX-r2-Ec-bioC_avg-signal461.352295
Spike_AFFX-r2-Bs-dap_avg-signal328.197601
Spike_AFFX-r2-Bs-dap_3-5-ratio9.049317
Spike_AFFX-r2-Ec-bioB_5_signal22.474264
ScaleMaskAll
BG2
VZ4
TGT100
Alpha10.040000
Alpha20.060000
SF0.970609
NF1.000000
HZ4
Tau0.015000

Slide: Rayson_647-19_upf3-1_NMD_3hr _cordycepin_Rep1_ATH1

Add to Slide Selection

Get the data!

Summary BioSource 1 Information Extraction / Labelling Protocol for Source 1 Information Slide Information Hybridisation Analysis Information for "Affymetrix MAS 5.0 Scaling Protocol" normalised data.

Summary Information

This slide has had 1 extract(s) hybridised to it (1-colour).

  • Extract 1 ("Rayson_647-19_upf3-1_NMD_3hr _cordycepin_Rep1_ATH1") was made from "Arabidopsis thaliana" and labelled using "Biotin Labelled cRNA using Affymetrix Enzo Kit".

The slide used was "Affymetrix ATH1 Arabidopsis Genome Array".

Click on the options above for more information about this slide and its preparation.

Hybridisation Protocol Information

Name of Protocol: Affymetrix Antibody Amplification Protocol
Solution:
BufferEDTA, NaCl, Tween20 according to Affymetrix protocol
Water
Blocking Agent:
Herring Sperm DNA(Source: Promega)100pg/ml
Acetylated BSA(Source: Sigma-Aldrich)0.1mg/ml
Wash Procedure:
Step 1(Source: Affymetrix)10 cycles of 2 mixes/cycle with Non-Stringent buffer
Step 2(Source: Affymetrix)4 cycles of 15 mixes/cycle with Stringent buffer
Step 3(Source: Affymetrix)Stain the probe array for 10 minutes in Streptavaidin Phycoerythrin (SAPE) solution at 25 C
Step 4(Source: Affymetrix)10 cycles of 4 mixes/cycle with Non-Stringent buffer
Step 5(Source: Affymetrix)Stain the probe array for 10 minutes in antibody solution
Step 6(Source: Affymetrix)Stain the probe array for 10 minutes in SAPE solution at 25 C
Step 7(Source: Affymetrix)15 cycles of 4mixes/cycle with Non-Stringent buffer at 30 C
Step 8(Source: Affymetrix)Hold at 25 C
Quantity Used:
Amount15 microgram Fragmented cRNA
Time: 16 hours
Concentration: 50 pg/ml
Volume: 200 microlitres
Temperature: 45 C
Other Information:
Spike(Source: Affymetrix)Control Oligo B2 to 50pM for landing lights
Spike(Source: Affymetrix)20X Eukaryotic Hybridisation Controls (bioB (1.5pM, bioC 5pM, bioD 25pM, cre 100pM)
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

BioSource 1 Information

BioSource Name: Rayson_647-19_upf3-1_NMD_3hr _cordycepin_Rep1_ATH1
Organism: Arabidopsis thaliana
Alias: upf3-1
Stock Code: N9900
Genetic Background: Col-0
Age: 3 weeks
Growth Conditions:
StratificationImbibed in the dark at 4 °C for 3 days.
ProtocolPlants were grown in a Sanyo MLR-352 H with additive humidity under a typical short day cycle (8hr light:16 hr dark) The temperature was maintained at 21-23°C and the humidity was maintained at 65 % RH. Stratified seeds were sown onto SHL compost at a density of 4 seed per cell (24 cell tray). Trays were covered in a propagator lid, which was removed 7 days after sowing. Compost was kept moist throughout.
Soil ConstituentsSinclair Potting Growth Medium (www.william-sinclair.co.uk)
Plant Spacing1 per cell
waterCompost was kept moist at all times
Temperature22 °C average, 23 °C day, 21 °C night
Humidity65 % average, 65 % day, 65 % night
MediumNone
Lighting(Source: Cool white fluorescent manufactured by SANYO. Intensity: 175µEinsteins. Wavelength: White)
Developmental Stage:
Growth Stage(Source: Paradigm Genetics)
Tissue: Aerial tissues
in vivo Treatment: Time = 3 hours Aerial tissues were harvested into incubation buffer (IB: 1 mM Pipes, 1 mM Sodium citrate, 1 mM Potassium chloride, 15 mM Sucrose). After 30 minutes in IB, all except the time (T) = 0 samples were transferred to IB with cordycepin (150 µg ml-1). The T = 0 samples were transferred to fresh IB without cordycepin. All flasks (including the T = 0) were transferred to a bell jar and vacuum infiltrated. Immediately post-infiltration was classed as T =0.
Additional Organism Information:
Sample DescriptionSoil-grown plant grown under short-day conditions.
Other Information:
Timecourse start procedureThe time-course started immediately after the vacuum infiltration.

Protocols for BioSource 1

Extraction Protocol Information

Name of Protocol: Qiagen / Trizol
Method:
ProtocolQiagen RNAeasy kit protocol
Type: total RNA
Amplification: Amplified during labelling process - see labelling protocol
Other Information:
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

Labelling Protocol Information

Name of Protocol: Biotin Labelled cRNA using Affymetrix Enzo Kit
Amount Labelled: Unknown
Label Name: Biotin
Labelling Method: In-vitro transcription method
Other Information:
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

Slide Information

Date Hybridised:

Array Design Information

Slide Design Name: Affymetrix ATH1 Arabidopsis Genome Array
Array Type: Affymetrix Gene Chip
Number of Spots on the Array: 22810

Scanning Protocol Information

Name: Affymetrix MAS 5.0 Standard Scanning
Width of Image Produced (Pixels): 4733
Height of Image Produced (Pixels): 4733
X Resolution (microns): 3
Y Resolution (microns): 3
Scanner Name: Agilent 2500A GeneArray Scanner
Software Name: Affymetrix Micro Array Suite 5.0

Image Analysis Information

Scaling Factor:0.511231
Name: Affymetrix MAS 5.0 Standard Image Analysis
Software Name: Affymetrix Microarray Analysis Suite 5.0

Normalisation Information

Strategy Name: Affymetrix MAS 5.0 Scaling Protocol
Algorithm: The top 2% and bottom 2% of signal intensities are excluded, then the mean is calculated. The original signal values are scaled such that the mean is made equal to 100.
Control Elements:
none
Normalisation Protocol Parameters:
Spike_AFFX-r2-P1-cre_3-5-ratio1.286346
Spike_AFFX-r2-Ec-bioB_3-5-ratio3.268257
Spike_AFFX-r2-Bs-dap_5_signal33.203674
NoiseAvg:3.62,Std:0.16,Min:3.1,Max:4.1
Control DirectionAntisense
Spike_AFFX-r2-Ec-bioB_5_detectionA
Spike_AFFX-r2-Ec-bioB_3_detectionP
Spike_AFFX-r2-Bs-thr_5_signal15.710994
#P12730
Spike_AFFX-r2-Bs-phe_M_signal29.821398
Corner-Avg:12288,Count:32
Spike_AFFX-r2-P1-cre_3_detectionP
Spike_AFFX-r2-Ec-bioB_M_signal15.721306
Spike_AFFX-r2-Ec-bioB_3_signal16.354534
Spike_AFFX-r2-Bs-lys_M_signal21.749485
Spike_AFFX-r2-P1-cre_3_signal4220.887695
Spike_AFFX-r2-Bs-lys_3-5-ratio1.728723
Spike_AFFX-r2-Bs-dap_M_signal171.613037
Spike_AFFX-r2-Bs-thr_M_detectionP
Probe Pair Threshold1
Spike_AFFX-r2-Bs-thr_3-5-ratio8.640942
Spike_AFFX-r2-Ec-bioB_avg-signal12.359965
Spike_AFFX-r2-Bs-thr_avg-signal69.266502
Spike_AFFX-r2-Bs-thr_5_detectionP
Spike_AFFX-r2-Bs-dap_3_detectionP
Spike_AFFX-r2-Ec-bioD_5_signal850.812927
Spike_AFFX-r2-Bs-phe_5_signal24.470747
Spike_AFFX-r2-P1-cre_5_detectionP
Spike_AFFX-r2-Ec-bioD_3_signal1257.885620
RawQ3.093527
Spike_AFFX-r2-Bs-lys_5_signal25.538490
Signal(A)5.590365
%A41.907059
Signal(All)152.665802
Corner+Avg:205,Count:32
Spike_AFFX-r2-Ec-bioD_5_detectionP
Spike_AFFX-r2-Ec-bioC_3_signal232.217926
Spike_AFFX-r2-Bs-lys_3_detectionP
Spike_AFFX-r2-Bs-phe_3_signal65.005020
Spike_AFFX-r2-Ec-bioD_avg-signal1054.349243
Spike_AFFX-r2-Bs-dap_5_detectionP
Spike_AFFX-r2-Bs-phe_5_detectionP
%P55.808857
Spike_AFFX-r2-Bs-lys_avg-signal30.478987
Spike_AFFX-r2-P1-cre_avg-signal3751.094727
Spike_AFFX-r2-Ec-bioB_M_detectionP
Spike_AFFX-r2-Bs-lys_M_detectionP
#Probe Sets Exceeding Probe Pair Threshold22810
%M2.284086
Spike_AFFX-r2-Bs-thr_3_signal135.757782
Spike_AFFX-r2-Ec-bioC_5_detectionP
Spike_AFFX-r2-Bs-phe_3_detectionP
Spike_AFFX-r2-Ec-bioC_3_detectionP
Spike_AFFX-r2-Bs-phe_avg-signal39.765720
Spike_AFFX-r2-Bs-dap_3_signal329.519104
Spike_AFFX-r2-Ec-bioD_3-5-ratio1.478451
Spike_AFFX-r2-Ec-bioD_3_detectionP
Spike_AFFX-r2-Ec-bioC_5_signal220.705826
#M521
Spike_AFFX-r2-Bs-lys_5_detectionP
Spike_AFFX-r2-Bs-lys_3_signal44.148987
Spike_AFFX-r2-Bs-dap_M_detectionP
Spike_AFFX-r2-Ec-bioC_3-5-ratio1.052160
Spike_AFFX-r2-Bs-thr_M_signal56.330734
Signal(P)268.593170
Spike_AFFX-r2-Bs-phe_3-5-ratio2.656438
Spike_AFFX-r2-Bs-thr_3_detectionP
Central-Avg:10914,Count:9
Spike_AFFX-r2-P1-cre_5_signal3281.301514
Spike_AFFX-r2-Bs-phe_M_detectionP
#A9559
Signal(M)18.574385
BackgroundAvg:77.29,Std:0.91,Min:75.3,Max:79.4
Spike_AFFX-r2-Ec-bioC_avg-signal226.461884
Spike_AFFX-r2-Bs-dap_avg-signal178.111938
Spike_AFFX-r2-Bs-dap_3-5-ratio9.924176
Spike_AFFX-r2-Ec-bioB_5_signal5.004055
ScaleMaskAll
BG2
VZ4
TGT100
Alpha10.040000
Alpha20.060000
SF0.511231
NF1.000000
HZ4
Tau0.015000

Slide: Rayson_647-18_upf3-1_NMD_1hr _cordycepin_Rep1_ATH1

Add to Slide Selection

Get the data!

Summary BioSource 1 Information Extraction / Labelling Protocol for Source 1 Information Slide Information Hybridisation Analysis Information for "Affymetrix MAS 5.0 Scaling Protocol" normalised data.

Summary Information

This slide has had 1 extract(s) hybridised to it (1-colour).

  • Extract 1 ("Rayson_647-18_upf3-1_NMD_1hr _cordycepin_Rep1_ATH1") was made from "Arabidopsis thaliana" and labelled using "Biotin Labelled cRNA using Affymetrix Enzo Kit".

The slide used was "Affymetrix ATH1 Arabidopsis Genome Array".

Click on the options above for more information about this slide and its preparation.

Hybridisation Protocol Information

Name of Protocol: Affymetrix Antibody Amplification Protocol
Solution:
BufferEDTA, NaCl, Tween20 according to Affymetrix protocol
Water
Blocking Agent:
Herring Sperm DNA(Source: Promega)100pg/ml
Acetylated BSA(Source: Sigma-Aldrich)0.1mg/ml
Wash Procedure:
Step 1(Source: Affymetrix)10 cycles of 2 mixes/cycle with Non-Stringent buffer
Step 2(Source: Affymetrix)4 cycles of 15 mixes/cycle with Stringent buffer
Step 3(Source: Affymetrix)Stain the probe array for 10 minutes in Streptavaidin Phycoerythrin (SAPE) solution at 25 C
Step 4(Source: Affymetrix)10 cycles of 4 mixes/cycle with Non-Stringent buffer
Step 5(Source: Affymetrix)Stain the probe array for 10 minutes in antibody solution
Step 6(Source: Affymetrix)Stain the probe array for 10 minutes in SAPE solution at 25 C
Step 7(Source: Affymetrix)15 cycles of 4mixes/cycle with Non-Stringent buffer at 30 C
Step 8(Source: Affymetrix)Hold at 25 C
Quantity Used:
Amount15 microgram Fragmented cRNA
Time: 16 hours
Concentration: 50 pg/ml
Volume: 200 microlitres
Temperature: 45 C
Other Information:
Spike(Source: Affymetrix)Control Oligo B2 to 50pM for landing lights
Spike(Source: Affymetrix)20X Eukaryotic Hybridisation Controls (bioB (1.5pM, bioC 5pM, bioD 25pM, cre 100pM)
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

BioSource 1 Information

BioSource Name: Rayson_647-18_upf3-1_NMD_1hr _cordycepin_Rep1_ATH1
Organism: Arabidopsis thaliana
Alias: upf3-1
Stock Code: N9900
Genetic Background: Col-0
Age: 3 weeks
Growth Conditions:
StratificationImbibed in the dark at 4 °C for 3 days.
ProtocolPlants were grown in a Sanyo MLR-352 H with additive humidity under a typical short day cycle (8hr light:16 hr dark) The temperature was maintained at 21-23°C and the humidity was maintained at 65 % RH. Stratified seeds were sown onto SHL compost at a density of 4 seed per cell (24 cell tray). Trays were covered in a propagator lid, which was removed 7 days after sowing. Compost was kept moist throughout.
Soil ConstituentsSinclair Potting Growth Medium (www.william-sinclair.co.uk)
Plant Spacing1 per cell
waterCompost was kept moist at all times
Temperature22 °C average, 23 °C day, 21 °C night
Humidity65 % average, 65 % day, 65 % night
MediumNone
Lighting(Source: Cool white fluorescent manufactured by SANYO. Intensity: 175µEinsteins. Wavelength: White)
Developmental Stage:
Growth Stage(Source: Paradigm Genetics)
Tissue: Aerial tissues
in vivo Treatment: Time = 1 hour Aerial tissues were harvested into incubation buffer (IB: 1 mM Pipes, 1 mM Sodium citrate, 1 mM Potassium chloride, 15 mM Sucrose). After 30 minutes in IB, all except the time (T) = 0 samples were transferred to IB with cordycepin (150 µg ml-1). The T = 0 samples were transferred to fresh IB without cordycepin. All flasks (including the T = 0) were transferred to a bell jar and vacuum infiltrated. Immediately post-infiltration was classed as T =0.
Additional Organism Information:
Sample DescriptionSoil-grown plant grown under short-day conditions.
Other Information:
Timecourse start procedureThe time-course started immediately after the vacuum infiltration.

Protocols for BioSource 1

Extraction Protocol Information

Name of Protocol: Qiagen / Trizol
Method:
ProtocolQiagen RNAeasy kit protocol
Type: total RNA
Amplification: Amplified during labelling process - see labelling protocol
Other Information:
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

Labelling Protocol Information

Name of Protocol: Biotin Labelled cRNA using Affymetrix Enzo Kit
Amount Labelled: Unknown
Label Name: Biotin
Labelling Method: In-vitro transcription method
Other Information:
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

Slide Information

Date Hybridised:

Array Design Information

Slide Design Name: Affymetrix ATH1 Arabidopsis Genome Array
Array Type: Affymetrix Gene Chip
Number of Spots on the Array: 22810

Scanning Protocol Information

Name: Affymetrix MAS 5.0 Standard Scanning
Width of Image Produced (Pixels): 4733
Height of Image Produced (Pixels): 4733
X Resolution (microns): 3
Y Resolution (microns): 3
Scanner Name: Agilent 2500A GeneArray Scanner
Software Name: Affymetrix Micro Array Suite 5.0

Image Analysis Information

Scaling Factor:0.547506
Name: Affymetrix MAS 5.0 Standard Image Analysis
Software Name: Affymetrix Microarray Analysis Suite 5.0

Normalisation Information

Strategy Name: Affymetrix MAS 5.0 Scaling Protocol
Algorithm: The top 2% and bottom 2% of signal intensities are excluded, then the mean is calculated. The original signal values are scaled such that the mean is made equal to 100.
Control Elements:
none
Normalisation Protocol Parameters:
Spike_AFFX-r2-P1-cre_3-5-ratio1.246212
Spike_AFFX-r2-Ec-bioB_3-5-ratio8.074231
Spike_AFFX-r2-Bs-dap_5_signal32.581112
NoiseAvg:3.67,Std:0.14,Min:3.4,Max:4.0
Control DirectionAntisense
Spike_AFFX-r2-Ec-bioB_5_detectionA
Spike_AFFX-r2-Ec-bioB_3_detectionA
Spike_AFFX-r2-Bs-thr_5_signal14.069494
#P13188
Spike_AFFX-r2-Bs-phe_M_signal24.297815
Corner-Avg:10403,Count:32
Spike_AFFX-r2-P1-cre_3_detectionP
Spike_AFFX-r2-Ec-bioB_M_signal8.651401
Spike_AFFX-r2-Ec-bioB_3_signal16.655083
Spike_AFFX-r2-Bs-lys_M_signal16.467916
Spike_AFFX-r2-P1-cre_3_signal3623.715088
Spike_AFFX-r2-Bs-lys_3-5-ratio3.138848
Spike_AFFX-r2-Bs-dap_M_signal161.936996
Spike_AFFX-r2-Bs-thr_M_detectionP
Probe Pair Threshold1
Spike_AFFX-r2-Bs-thr_3-5-ratio6.007506
Spike_AFFX-r2-Ec-bioB_avg-signal9.123076
Spike_AFFX-r2-Bs-thr_avg-signal49.327045
Spike_AFFX-r2-Bs-thr_5_detectionP
Spike_AFFX-r2-Bs-dap_3_detectionP
Spike_AFFX-r2-Ec-bioD_5_signal648.906860
Spike_AFFX-r2-Bs-phe_5_signal20.752604
Spike_AFFX-r2-P1-cre_5_detectionP
Spike_AFFX-r2-Ec-bioD_3_signal1017.007324
RawQ3.214193
Spike_AFFX-r2-Bs-lys_5_signal16.838753
Signal(A)5.458451
%A40.109600
Signal(All)146.023239
Corner+Avg:173,Count:32
Spike_AFFX-r2-Ec-bioD_5_detectionP
Spike_AFFX-r2-Ec-bioC_3_signal198.459793
Spike_AFFX-r2-Bs-lys_3_detectionP
Spike_AFFX-r2-Bs-phe_3_signal49.884201
Spike_AFFX-r2-Ec-bioD_avg-signal832.957092
Spike_AFFX-r2-Bs-dap_5_detectionP
Spike_AFFX-r2-Bs-phe_5_detectionP
%P57.816746
Spike_AFFX-r2-Bs-lys_avg-signal28.720320
Spike_AFFX-r2-P1-cre_avg-signal3265.749023
Spike_AFFX-r2-Ec-bioB_M_detectionP
Spike_AFFX-r2-Bs-lys_M_detectionP
#Probe Sets Exceeding Probe Pair Threshold22810
%M2.073652
Spike_AFFX-r2-Bs-thr_3_signal84.522575
Spike_AFFX-r2-Ec-bioC_5_detectionP
Spike_AFFX-r2-Bs-phe_3_detectionP
Spike_AFFX-r2-Ec-bioC_3_detectionP
Spike_AFFX-r2-Bs-phe_avg-signal31.644873
Spike_AFFX-r2-Bs-dap_3_signal246.760590
Spike_AFFX-r2-Ec-bioD_3-5-ratio1.567262
Spike_AFFX-r2-Ec-bioD_3_detectionP
Spike_AFFX-r2-Ec-bioC_5_signal195.386597
#M473
Spike_AFFX-r2-Bs-lys_5_detectionP
Spike_AFFX-r2-Bs-lys_3_signal52.854290
Spike_AFFX-r2-Bs-dap_M_detectionP
Spike_AFFX-r2-Ec-bioC_3-5-ratio1.015729
Spike_AFFX-r2-Bs-thr_M_signal49.389084
Signal(P)248.101776
Spike_AFFX-r2-Bs-phe_3-5-ratio2.403756
Spike_AFFX-r2-Bs-thr_3_detectionP
Central-Avg:9493,Count:9
Spike_AFFX-r2-P1-cre_5_signal2907.783203
Spike_AFFX-r2-Bs-phe_M_detectionP
#A9149
Signal(M)18.783005
BackgroundAvg:81.77,Std:0.77,Min:79.8,Max:83.6
Spike_AFFX-r2-Ec-bioC_avg-signal196.923187
Spike_AFFX-r2-Bs-dap_avg-signal147.092896
Spike_AFFX-r2-Bs-dap_3-5-ratio7.573731
Spike_AFFX-r2-Ec-bioB_5_signal2.062745
ScaleMaskAll
BG2
VZ4
TGT100
Alpha10.040000
Alpha20.060000
SF0.547506
NF1.000000
HZ4
Tau0.015000

Slide: Rayson_647-17_upf3-1_NMD_0hr_cordycepin_Rep1_ATH1

Add to Slide Selection

Get the data!

Summary BioSource 1 Information Extraction / Labelling Protocol for Source 1 Information Slide Information Hybridisation Analysis Information for "Affymetrix MAS 5.0 Scaling Protocol" normalised data.

Summary Information

This slide has had 1 extract(s) hybridised to it (1-colour).

  • Extract 1 ("Rayson_647-17_upf3-1_NMD_0hr_cordycepin_Rep1_ATH1") was made from "Arabidopsis thaliana" and labelled using "Biotin Labelled cRNA using Affymetrix Enzo Kit".

The slide used was "Affymetrix ATH1 Arabidopsis Genome Array".

Click on the options above for more information about this slide and its preparation.

Hybridisation Protocol Information

Name of Protocol: Affymetrix Antibody Amplification Protocol
Solution:
BufferEDTA, NaCl, Tween20 according to Affymetrix protocol
Water
Blocking Agent:
Herring Sperm DNA(Source: Promega)100pg/ml
Acetylated BSA(Source: Sigma-Aldrich)0.1mg/ml
Wash Procedure:
Step 1(Source: Affymetrix)10 cycles of 2 mixes/cycle with Non-Stringent buffer
Step 2(Source: Affymetrix)4 cycles of 15 mixes/cycle with Stringent buffer
Step 3(Source: Affymetrix)Stain the probe array for 10 minutes in Streptavaidin Phycoerythrin (SAPE) solution at 25 C
Step 4(Source: Affymetrix)10 cycles of 4 mixes/cycle with Non-Stringent buffer
Step 5(Source: Affymetrix)Stain the probe array for 10 minutes in antibody solution
Step 6(Source: Affymetrix)Stain the probe array for 10 minutes in SAPE solution at 25 C
Step 7(Source: Affymetrix)15 cycles of 4mixes/cycle with Non-Stringent buffer at 30 C
Step 8(Source: Affymetrix)Hold at 25 C
Quantity Used:
Amount15 microgram Fragmented cRNA
Time: 16 hours
Concentration: 50 pg/ml
Volume: 200 microlitres
Temperature: 45 C
Other Information:
Spike(Source: Affymetrix)Control Oligo B2 to 50pM for landing lights
Spike(Source: Affymetrix)20X Eukaryotic Hybridisation Controls (bioB (1.5pM, bioC 5pM, bioD 25pM, cre 100pM)
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

BioSource 1 Information

BioSource Name: Rayson_647-17_upf3-1_NMD_0hr_cordycepin_Rep1_ATH1
Organism: Arabidopsis thaliana
Alias: upf3-1
Stock Code: N9900
Genetic Background: Col-0
Age: 3 weeks
Growth Conditions:
StratificationImbibed in the dark at 4 °C for 3 days.
ProtocolPlants were grown in a Sanyo MLR-352 H with additive humidity under a typical short day cycle (8hr light:16 hr dark) The temperature was maintained at 21-23°C and the humidity was maintained at 65 % RH. Stratified seeds were sown onto SHL compost at a density of 4 seed per cell (24 cell tray). Trays were covered in a propagator lid, which was removed 7 days after sowing. Compost was kept moist throughout.
Soil ConstituentsSinclair Potting Growth Medium (www.william-sinclair.co.uk)
Plant Spacing1 per cell
waterCompost was kept moist at all times
Temperature22 °C average, 23 °C day, 21 °C night
Humidity65 % average, 65 % day, 65 % night
MediumNone
Lighting(Source: Cool white fluorescent manufactured by SANYO. Intensity: 175µEinsteins. Wavelength: White)
Developmental Stage:
Growth Stage(Source: Paradigm Genetics)
Tissue: Aerial tissues
in vivo Treatment: Time = 0 Aerial tissues were harvested into incubation buffer (IB: 1 mM Pipes, 1 mM Sodium citrate, 1 mM Potassium chloride, 15 mM Sucrose). After 30 minutes in IB, all except the time (T) = 0 samples were transferred to IB with cordycepin (150 µg ml-1). The T = 0 samples were transferred to fresh IB without cordycepin. All flasks (including the T = 0) were transferred to a bell jar and vacuum infiltrated. Immediately post-infiltration was classed as T =0.
Additional Organism Information:
Sample DescriptionSoil-grown plant grown under short-day conditions.
Other Information:
Timecourse start procedureThe time-course started immediately after the vacuum infiltration.

Protocols for BioSource 1

Extraction Protocol Information

Name of Protocol: Qiagen / Trizol
Method:
ProtocolQiagen RNAeasy kit protocol
Type: total RNA
Amplification: Amplified during labelling process - see labelling protocol
Other Information:
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

Labelling Protocol Information

Name of Protocol: Biotin Labelled cRNA using Affymetrix Enzo Kit
Amount Labelled: Unknown
Label Name: Biotin
Labelling Method: In-vitro transcription method
Other Information:
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

Slide Information

Date Hybridised:

Array Design Information

Slide Design Name: Affymetrix ATH1 Arabidopsis Genome Array
Array Type: Affymetrix Gene Chip
Number of Spots on the Array: 22810

Scanning Protocol Information

Name: Affymetrix MAS 5.0 Standard Scanning
Width of Image Produced (Pixels): 4733
Height of Image Produced (Pixels): 4733
X Resolution (microns): 3
Y Resolution (microns): 3
Scanner Name: Agilent 2500A GeneArray Scanner
Software Name: Affymetrix Micro Array Suite 5.0

Image Analysis Information

Scaling Factor:0.404873
Name: Affymetrix MAS 5.0 Standard Image Analysis
Software Name: Affymetrix Microarray Analysis Suite 5.0

Normalisation Information

Strategy Name: Affymetrix MAS 5.0 Scaling Protocol
Algorithm: The top 2% and bottom 2% of signal intensities are excluded, then the mean is calculated. The original signal values are scaled such that the mean is made equal to 100.
Control Elements:
none
Normalisation Protocol Parameters:
Spike_AFFX-r2-P1-cre_3-5-ratio1.267448
Spike_AFFX-r2-Ec-bioB_3-5-ratio2.243719
Spike_AFFX-r2-Bs-dap_5_signal25.800974
NoiseAvg:3.74,Std:0.08,Min:3.5,Max:4.0
Control DirectionAntisense
Spike_AFFX-r2-Ec-bioB_5_detectionA
Spike_AFFX-r2-Ec-bioB_3_detectionM
Spike_AFFX-r2-Bs-thr_5_signal7.523598
#P14286
Spike_AFFX-r2-Bs-phe_M_signal14.083508
Corner-Avg:12227,Count:32
Spike_AFFX-r2-P1-cre_3_detectionP
Spike_AFFX-r2-Ec-bioB_M_signal9.662626
Spike_AFFX-r2-Ec-bioB_3_signal13.165941
Spike_AFFX-r2-Bs-lys_M_signal14.383219
Spike_AFFX-r2-P1-cre_3_signal3792.766846
Spike_AFFX-r2-Bs-lys_3-5-ratio1.983357
Spike_AFFX-r2-Bs-dap_M_signal92.531853
Spike_AFFX-r2-Bs-thr_M_detectionP
Probe Pair Threshold1
Spike_AFFX-r2-Bs-thr_3-5-ratio8.807486
Spike_AFFX-r2-Ec-bioB_avg-signal9.565492
Spike_AFFX-r2-Bs-thr_avg-signal32.936939
Spike_AFFX-r2-Bs-thr_5_detectionA
Spike_AFFX-r2-Bs-dap_3_detectionP
Spike_AFFX-r2-Ec-bioD_5_signal768.325012
Spike_AFFX-r2-Bs-phe_5_signal12.854889
Spike_AFFX-r2-P1-cre_5_detectionP
Spike_AFFX-r2-Ec-bioD_3_signal1011.621643
RawQ3.026045
Spike_AFFX-r2-Bs-lys_5_signal12.730353
Signal(A)3.843831
%A35.716789
Signal(All)138.055710
Corner+Avg:210,Count:32
Spike_AFFX-r2-Ec-bioD_5_detectionP
Spike_AFFX-r2-Ec-bioC_3_signal220.722305
Spike_AFFX-r2-Bs-lys_3_detectionP
Spike_AFFX-r2-Bs-phe_3_signal36.974167
Spike_AFFX-r2-Ec-bioD_avg-signal889.973328
Spike_AFFX-r2-Bs-dap_5_detectionP
Spike_AFFX-r2-Bs-phe_5_detectionP
%P62.630424
Spike_AFFX-r2-Bs-lys_avg-signal17.454138
Spike_AFFX-r2-P1-cre_avg-signal3392.605225
Spike_AFFX-r2-Ec-bioB_M_detectionP
Spike_AFFX-r2-Bs-lys_M_detectionP
#Probe Sets Exceeding Probe Pair Threshold22810
%M1.652784
Spike_AFFX-r2-Bs-thr_3_signal66.263985
Spike_AFFX-r2-Ec-bioC_5_detectionP
Spike_AFFX-r2-Bs-phe_3_detectionP
Spike_AFFX-r2-Ec-bioC_3_detectionP
Spike_AFFX-r2-Bs-phe_avg-signal21.304188
Spike_AFFX-r2-Bs-dap_3_signal197.923248
Spike_AFFX-r2-Ec-bioD_3-5-ratio1.316658
Spike_AFFX-r2-Ec-bioD_3_detectionP
Spike_AFFX-r2-Ec-bioC_5_signal198.470093
#M377
Spike_AFFX-r2-Bs-lys_5_detectionP
Spike_AFFX-r2-Bs-lys_3_signal25.248838
Spike_AFFX-r2-Bs-dap_M_detectionP
Spike_AFFX-r2-Ec-bioC_3-5-ratio1.112119
Spike_AFFX-r2-Bs-thr_M_signal25.023232
Signal(P)217.830551
Spike_AFFX-r2-Bs-phe_3-5-ratio2.876273
Spike_AFFX-r2-Bs-thr_3_detectionP
Central-Avg:11252,Count:9
Spike_AFFX-r2-P1-cre_5_signal2992.443604
Spike_AFFX-r2-Bs-phe_M_detectionP
#A8147
Signal(M)15.405325
BackgroundAvg:76.26,Std:0.94,Min:73.7,Max:77.8
Spike_AFFX-r2-Ec-bioC_avg-signal209.596191
Spike_AFFX-r2-Bs-dap_avg-signal105.418694
Spike_AFFX-r2-Bs-dap_3-5-ratio7.671154
Spike_AFFX-r2-Ec-bioB_5_signal5.867909
ScaleMaskAll
BG2
VZ4
TGT100
Alpha10.040000
Alpha20.060000
SF0.404873
NF1.000000
HZ4
Tau0.015000

Slide: Rayson_647-16_UPF2(6-11E5)_NMD_6hr _cordycepin_Rep1_ATH1

Add to Slide Selection

Get the data!

Summary BioSource 1 Information Extraction / Labelling Protocol for Source 1 Information Slide Information Hybridisation Analysis Information for "Affymetrix MAS 5.0 Scaling Protocol" normalised data.

Summary Information

This slide has had 1 extract(s) hybridised to it (1-colour).

  • Extract 1 ("Rayson_647-16_UPF2(6-11E5)_NMD_6hr _cordycepin_Rep1_ATH1") was made from "Arabidopsis thaliana" and labelled using "Biotin Labelled cRNA using Affymetrix Enzo Kit".

The slide used was "Affymetrix ATH1 Arabidopsis Genome Array".

Click on the options above for more information about this slide and its preparation.

Hybridisation Protocol Information

Name of Protocol: Affymetrix Antibody Amplification Protocol
Solution:
BufferEDTA, NaCl, Tween20 according to Affymetrix protocol
Water
Blocking Agent:
Herring Sperm DNA(Source: Promega)100pg/ml
Acetylated BSA(Source: Sigma-Aldrich)0.1mg/ml
Wash Procedure:
Step 1(Source: Affymetrix)10 cycles of 2 mixes/cycle with Non-Stringent buffer
Step 2(Source: Affymetrix)4 cycles of 15 mixes/cycle with Stringent buffer
Step 3(Source: Affymetrix)Stain the probe array for 10 minutes in Streptavaidin Phycoerythrin (SAPE) solution at 25 C
Step 4(Source: Affymetrix)10 cycles of 4 mixes/cycle with Non-Stringent buffer
Step 5(Source: Affymetrix)Stain the probe array for 10 minutes in antibody solution
Step 6(Source: Affymetrix)Stain the probe array for 10 minutes in SAPE solution at 25 C
Step 7(Source: Affymetrix)15 cycles of 4mixes/cycle with Non-Stringent buffer at 30 C
Step 8(Source: Affymetrix)Hold at 25 C
Quantity Used:
Amount15 microgram Fragmented cRNA
Time: 16 hours
Concentration: 50 pg/ml
Volume: 200 microlitres
Temperature: 45 C
Other Information:
Spike(Source: Affymetrix)Control Oligo B2 to 50pM for landing lights
Spike(Source: Affymetrix)20X Eukaryotic Hybridisation Controls (bioB (1.5pM, bioC 5pM, bioD 25pM, cre 100pM)
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

BioSource 1 Information

BioSource Name: Rayson_647-16_UPF2(6-11E5)_NMD_6hr _cordycepin_Rep1_ATH1
Organism: Arabidopsis thaliana
Alias: 6-11E5
Stock Code:
Age: 3 weeks
Growth Conditions:
StratificationImbibed in the dark at 4 °C for 3 days.
ProtocolPlants were grown in a Sanyo MLR-352 H with additive humidity under a typical short day cycle (8hr light:16 hr dark) The temperature was maintained at 21-23°C and the humidity was maintained at 65 % RH. Stratified seeds were sown onto SHL compost at a density of 4 seed per cell (24 cell tray). Trays were covered in a propagator lid, which was removed 7 days after sowing. Compost was kept moist throughout.
Soil ConstituentsSinclair Potting Growth Medium (www.william-sinclair.co.uk)
Plant Spacing1 per cell
waterCompost was kept moist at all times
Temperature22 °C average, 23 °C day, 21 °C night
Humidity65 % average, 65 % day, 65 % night
MediumNone
Lighting(Source: Cool white fluorescent manufactured by SANYO. Intensity: 175µEinsteins. Wavelength: White)
Developmental Stage:
Growth Stage(Source: Paradigm Genetics)
Genetic Variation: induced mutation mutation in At2g39260
Tissue: Aerial tissues
in vivo Treatment: Time = 6 hours Aerial tissues were harvested into incubation buffer (IB: 1 mM Pipes, 1 mM Sodium citrate, 1 mM Potassium chloride, 15 mM Sucrose). After 30 minutes in IB, all except the time (T) = 0 samples were transferred to IB with cordycepin (150 µg ml-1). The T = 0 samples were transferred to fresh IB without cordycepin. All flasks (including the T = 0) were transferred to a bell jar and vacuum infiltrated. Immediately post-infiltration was classed as T =0.
Additional Organism Information:
Sample DescriptionSoil-grown plant grown under short-day conditions.
Other Information:
Timecourse start procedureThe time-course started immediately after the vacuum infiltration.

Protocols for BioSource 1

Extraction Protocol Information

Name of Protocol: Qiagen / Trizol
Method:
ProtocolQiagen RNAeasy kit protocol
Type: total RNA
Amplification: Amplified during labelling process - see labelling protocol
Other Information:
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

Labelling Protocol Information

Name of Protocol: Biotin Labelled cRNA using Affymetrix Enzo Kit
Amount Labelled: Unknown
Label Name: Biotin
Labelling Method: In-vitro transcription method
Other Information:
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

Slide Information

Date Hybridised:

Array Design Information

Slide Design Name: Affymetrix ATH1 Arabidopsis Genome Array
Array Type: Affymetrix Gene Chip
Number of Spots on the Array: 22810

Scanning Protocol Information

Name: Affymetrix MAS 5.0 Standard Scanning
Width of Image Produced (Pixels): 4733
Height of Image Produced (Pixels): 4733
X Resolution (microns): 3
Y Resolution (microns): 3
Scanner Name: Agilent 2500A GeneArray Scanner
Software Name: Affymetrix Micro Array Suite 5.0

Image Analysis Information

Scaling Factor:0.842575
Name: Affymetrix MAS 5.0 Standard Image Analysis
Software Name: Affymetrix Microarray Analysis Suite 5.0

Normalisation Information

Strategy Name: Affymetrix MAS 5.0 Scaling Protocol
Algorithm: The top 2% and bottom 2% of signal intensities are excluded, then the mean is calculated. The original signal values are scaled such that the mean is made equal to 100.
Control Elements:
none
Normalisation Protocol Parameters:
Spike_AFFX-r2-P1-cre_3-5-ratio1.274092
Spike_AFFX-r2-Ec-bioB_3-5-ratio0.995721
Spike_AFFX-r2-Bs-dap_5_signal79.141457
NoiseAvg:3.14,Std:0.13,Min:2.9,Max:3.6
Control DirectionAntisense
Spike_AFFX-r2-Ec-bioB_5_detectionA
Spike_AFFX-r2-Ec-bioB_3_detectionP
Spike_AFFX-r2-Bs-thr_5_signal49.084309
#P11593
Spike_AFFX-r2-Bs-phe_M_signal56.328220
Corner-Avg:10709,Count:32
Spike_AFFX-r2-P1-cre_3_detectionP
Spike_AFFX-r2-Ec-bioB_M_signal8.320117
Spike_AFFX-r2-Ec-bioB_3_signal14.449749
Spike_AFFX-r2-Bs-lys_M_signal38.333565
Spike_AFFX-r2-P1-cre_3_signal5628.213379
Spike_AFFX-r2-Bs-lys_3-5-ratio1.369795
Spike_AFFX-r2-Bs-dap_M_signal343.519073
Spike_AFFX-r2-Bs-thr_M_detectionP
Probe Pair Threshold1
Spike_AFFX-r2-Bs-thr_3-5-ratio5.184030
Spike_AFFX-r2-Ec-bioB_avg-signal12.427238
Spike_AFFX-r2-Bs-thr_avg-signal142.395187
Spike_AFFX-r2-Bs-thr_5_detectionP
Spike_AFFX-r2-Bs-dap_3_detectionP
Spike_AFFX-r2-Ec-bioD_5_signal1046.867920
Spike_AFFX-r2-Bs-phe_5_signal50.663548
Spike_AFFX-r2-P1-cre_5_detectionP
Spike_AFFX-r2-Ec-bioD_3_signal1492.152344
RawQ2.877287
Spike_AFFX-r2-Bs-lys_5_signal50.788303
Signal(A)6.853674
%A46.896099
Signal(All)159.298111
Corner+Avg:192,Count:32
Spike_AFFX-r2-Ec-bioD_5_detectionP
Spike_AFFX-r2-Ec-bioC_3_signal290.730194
Spike_AFFX-r2-Bs-lys_3_detectionP
Spike_AFFX-r2-Bs-phe_3_signal82.841629
Spike_AFFX-r2-Ec-bioD_avg-signal1269.510132
Spike_AFFX-r2-Bs-dap_5_detectionP
Spike_AFFX-r2-Bs-phe_5_detectionP
%P50.824200
Spike_AFFX-r2-Bs-lys_avg-signal52.897137
Spike_AFFX-r2-P1-cre_avg-signal5022.822266
Spike_AFFX-r2-Ec-bioB_M_detectionA
Spike_AFFX-r2-Bs-lys_M_detectionP
#Probe Sets Exceeding Probe Pair Threshold22810
%M2.279702
Spike_AFFX-r2-Bs-thr_3_signal254.454514
Spike_AFFX-r2-Ec-bioC_5_detectionP
Spike_AFFX-r2-Bs-phe_3_detectionP
Spike_AFFX-r2-Ec-bioC_3_detectionP
Spike_AFFX-r2-Bs-phe_avg-signal63.277798
Spike_AFFX-r2-Bs-dap_3_signal548.298401
Spike_AFFX-r2-Ec-bioD_3-5-ratio1.425349
Spike_AFFX-r2-Ec-bioD_3_detectionP
Spike_AFFX-r2-Ec-bioC_5_signal282.681671
#M520
Spike_AFFX-r2-Bs-lys_5_detectionP
Spike_AFFX-r2-Bs-lys_3_signal69.569542
Spike_AFFX-r2-Bs-dap_M_detectionP
Spike_AFFX-r2-Ec-bioC_3-5-ratio1.028472
Spike_AFFX-r2-Bs-thr_M_signal123.646706
Signal(P)306.057678
Spike_AFFX-r2-Bs-phe_3-5-ratio1.635133
Spike_AFFX-r2-Bs-thr_3_detectionP
Central-Avg:8762,Count:9
Spike_AFFX-r2-P1-cre_5_signal4417.431641
Spike_AFFX-r2-Bs-phe_M_detectionP
#A10697
Signal(M)23.365051
BackgroundAvg:72.71,Std:0.81,Min:70.9,Max:76.1
Spike_AFFX-r2-Ec-bioC_avg-signal286.705933
Spike_AFFX-r2-Bs-dap_avg-signal323.652985
Spike_AFFX-r2-Bs-dap_3-5-ratio6.928081
Spike_AFFX-r2-Ec-bioB_5_signal14.511846
ScaleMaskAll
BG2
VZ4
TGT100
Alpha10.040000
Alpha20.060000
SF0.842575
NF1.000000
HZ4
Tau0.015000

Slide: Rayson_647-12_upf1-5_NMD_6hr_cordycepin_Rep1_ATH1

Add to Slide Selection

Get the data!

Summary BioSource 1 Information Extraction / Labelling Protocol for Source 1 Information Slide Information Hybridisation Analysis Information for "Affymetrix MAS 5.0 Scaling Protocol" normalised data.

Summary Information

This slide has had 1 extract(s) hybridised to it (1-colour).

  • Extract 1 ("Rayson_647-12_upf1-5_NMD_6hr_cordycepin_Rep1_ATH1") was made from "Arabidopsis thaliana" and labelled using "Biotin Labelled cRNA using Affymetrix Enzo Kit".

The slide used was "Affymetrix ATH1 Arabidopsis Genome Array".

Click on the options above for more information about this slide and its preparation.

Hybridisation Protocol Information

Name of Protocol: Affymetrix Antibody Amplification Protocol
Solution:
BufferEDTA, NaCl, Tween20 according to Affymetrix protocol
Water
Blocking Agent:
Herring Sperm DNA(Source: Promega)100pg/ml
Acetylated BSA(Source: Sigma-Aldrich)0.1mg/ml
Wash Procedure:
Step 1(Source: Affymetrix)10 cycles of 2 mixes/cycle with Non-Stringent buffer
Step 2(Source: Affymetrix)4 cycles of 15 mixes/cycle with Stringent buffer
Step 3(Source: Affymetrix)Stain the probe array for 10 minutes in Streptavaidin Phycoerythrin (SAPE) solution at 25 C
Step 4(Source: Affymetrix)10 cycles of 4 mixes/cycle with Non-Stringent buffer
Step 5(Source: Affymetrix)Stain the probe array for 10 minutes in antibody solution
Step 6(Source: Affymetrix)Stain the probe array for 10 minutes in SAPE solution at 25 C
Step 7(Source: Affymetrix)15 cycles of 4mixes/cycle with Non-Stringent buffer at 30 C
Step 8(Source: Affymetrix)Hold at 25 C
Quantity Used:
Amount15 microgram Fragmented cRNA
Time: 16 hours
Concentration: 50 pg/ml
Volume: 200 microlitres
Temperature: 45 C
Other Information:
Spike(Source: Affymetrix)Control Oligo B2 to 50pM for landing lights
Spike(Source: Affymetrix)20X Eukaryotic Hybridisation Controls (bioB (1.5pM, bioC 5pM, bioD 25pM, cre 100pM)
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

BioSource 1 Information

BioSource Name: Rayson_647-12_upf1-5_NMD_6hr_cordycepin_Rep1_ATH1
Organism: Arabidopsis thaliana
Alias: upf1-5
Stock Code: N9902
Genetic Background: Col-0
Age: 3 weeks
Growth Conditions:
StratificationImbibed in the dark at 4 °C for 3 days.
ProtocolPlants were grown in a Sanyo MLR-352 H with additive humidity under a typical short day cycle (8hr light:16 hr dark) The temperature was maintained at 21-23°C and the humidity was maintained at 65 % RH. Stratified seeds were sown onto SHL compost at a density of 4 seed per cell (24 cell tray). Trays were covered in a propagator lid, which was removed 7 days after sowing. Compost was kept moist throughout.
Soil ConstituentsSinclair Potting Growth Medium (www.william-sinclair.co.uk)
Plant Spacing1 per cell
waterCompost was kept moist at all times
Temperature22 °C average, 23 °C day, 21 °C night
Humidity65 % average, 65 % day, 65 % night
MediumNone
Lighting(Source: Cool white fluorescent manufactured by SANYO. Intensity: 175µEinsteins. Wavelength: White)
Developmental Stage:
Growth Stage(Source: Paradigm Genetics)
Tissue: Aerial tissues
in vivo Treatment: Time = 6 hours Aerial tissues were harvested into incubation buffer (IB: 1 mM Pipes, 1 mM Sodium citrate, 1 mM Potassium chloride, 15 mM Sucrose). After 30 minutes in IB, all except the time (T) = 0 samples were transferred to IB with cordycepin (150 µg ml-1). The T = 0 samples were transferred to fresh IB without cordycepin. All flasks (including the T = 0) were transferred to a bell jar and vacuum infiltrated. Immediately post-infiltration was classed as T =0.
Additional Organism Information:
Sample DescriptionSoil-grown plant grown under short-day conditions.
Other Information:
Timecourse start procedureThe time-course started immediately after the vacuum infiltration.

Protocols for BioSource 1

Extraction Protocol Information

Name of Protocol: Qiagen / Trizol
Method:
ProtocolQiagen RNAeasy kit protocol
Type: total RNA
Amplification: Amplified during labelling process - see labelling protocol
Other Information:
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

Labelling Protocol Information

Name of Protocol: Biotin Labelled cRNA using Affymetrix Enzo Kit
Amount Labelled: Unknown
Label Name: Biotin
Labelling Method: In-vitro transcription method
Other Information:
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

Slide Information

Date Hybridised:

Array Design Information

Slide Design Name: Affymetrix ATH1 Arabidopsis Genome Array
Array Type: Affymetrix Gene Chip
Number of Spots on the Array: 22810

Scanning Protocol Information

Name: Affymetrix MAS 5.0 Standard Scanning
Width of Image Produced (Pixels): 4733
Height of Image Produced (Pixels): 4733
X Resolution (microns): 3
Y Resolution (microns): 3
Scanner Name: Agilent 2500A GeneArray Scanner
Software Name: Affymetrix Micro Array Suite 5.0

Image Analysis Information

Scaling Factor:1.203084
Name: Affymetrix MAS 5.0 Standard Image Analysis
Software Name: Affymetrix Microarray Analysis Suite 5.0

Normalisation Information

Strategy Name: Affymetrix MAS 5.0 Scaling Protocol
Algorithm: The top 2% and bottom 2% of signal intensities are excluded, then the mean is calculated. The original signal values are scaled such that the mean is made equal to 100.
Control Elements:
none
Normalisation Protocol Parameters:
Spike_AFFX-r2-P1-cre_3-5-ratio1.091698
Spike_AFFX-r2-Ec-bioB_3-5-ratio1.527381
Spike_AFFX-r2-Bs-dap_5_signal84.565025
NoiseAvg:2.65,Std:0.10,Min:2.3,Max:2.9
Control DirectionAntisense
Spike_AFFX-r2-Ec-bioB_5_detectionP
Spike_AFFX-r2-Ec-bioB_3_detectionP
Spike_AFFX-r2-Bs-thr_5_signal36.671474
#P10994
Spike_AFFX-r2-Bs-phe_M_signal57.876846
Corner-Avg:11510,Count:32
Spike_AFFX-r2-P1-cre_3_detectionP
Spike_AFFX-r2-Ec-bioB_M_signal19.895943
Spike_AFFX-r2-Ec-bioB_3_signal26.362345
Spike_AFFX-r2-Bs-lys_M_signal41.067684
Spike_AFFX-r2-P1-cre_3_signal8695.797852
Spike_AFFX-r2-Bs-lys_3-5-ratio1.272921
Spike_AFFX-r2-Bs-dap_M_signal298.008362
Spike_AFFX-r2-Bs-thr_M_detectionP
Probe Pair Threshold1
Spike_AFFX-r2-Bs-thr_3-5-ratio7.215606
Spike_AFFX-r2-Ec-bioB_avg-signal21.172709
Spike_AFFX-r2-Bs-thr_avg-signal139.964035
Spike_AFFX-r2-Bs-thr_5_detectionP
Spike_AFFX-r2-Bs-dap_3_detectionP
Spike_AFFX-r2-Ec-bioD_5_signal2415.627441
Spike_AFFX-r2-Bs-phe_5_signal32.147186
Spike_AFFX-r2-P1-cre_5_detectionP
Spike_AFFX-r2-Ec-bioD_3_signal2972.355469
RawQ2.621779
Spike_AFFX-r2-Bs-lys_5_signal29.668760
Signal(A)7.667392
%A49.368698
Signal(All)169.816452
Corner+Avg:196,Count:32
Spike_AFFX-r2-Ec-bioD_5_detectionP
Spike_AFFX-r2-Ec-bioC_3_signal597.460022
Spike_AFFX-r2-Bs-lys_3_detectionP
Spike_AFFX-r2-Bs-phe_3_signal108.708549
Spike_AFFX-r2-Ec-bioD_avg-signal2693.991455
Spike_AFFX-r2-Bs-dap_5_detectionP
Spike_AFFX-r2-Bs-phe_5_detectionP
%P48.198158
Spike_AFFX-r2-Bs-lys_avg-signal36.167484
Spike_AFFX-r2-P1-cre_avg-signal8330.593750
Spike_AFFX-r2-Ec-bioB_M_detectionP
Spike_AFFX-r2-Bs-lys_M_detectionP
#Probe Sets Exceeding Probe Pair Threshold22810
%M2.433143
Spike_AFFX-r2-Bs-thr_3_signal264.606903
Spike_AFFX-r2-Ec-bioC_5_detectionP
Spike_AFFX-r2-Bs-phe_3_detectionP
Spike_AFFX-r2-Ec-bioC_3_detectionP
Spike_AFFX-r2-Bs-phe_avg-signal66.244194
Spike_AFFX-r2-Bs-dap_3_signal445.278534
Spike_AFFX-r2-Ec-bioD_3-5-ratio1.230469
Spike_AFFX-r2-Ec-bioD_3_detectionP
Spike_AFFX-r2-Ec-bioC_5_signal527.358765
#M555
Spike_AFFX-r2-Bs-lys_5_detectionP
Spike_AFFX-r2-Bs-lys_3_signal37.766003
Spike_AFFX-r2-Bs-dap_M_detectionP
Spike_AFFX-r2-Ec-bioC_3-5-ratio1.132929
Spike_AFFX-r2-Bs-thr_M_signal118.613708
Signal(P)343.216583
Spike_AFFX-r2-Bs-phe_3-5-ratio3.381588
Spike_AFFX-r2-Bs-thr_3_detectionP
Central-Avg:9804,Count:9
Spike_AFFX-r2-P1-cre_5_signal7965.389160
Spike_AFFX-r2-Bs-phe_M_detectionP
#A11261
Signal(M)24.950340
BackgroundAvg:64.40,Std:0.50,Min:63.6,Max:65.9
Spike_AFFX-r2-Ec-bioC_avg-signal562.409424
Spike_AFFX-r2-Bs-dap_avg-signal275.950623
Spike_AFFX-r2-Bs-dap_3-5-ratio5.265516
Spike_AFFX-r2-Ec-bioB_5_signal17.259838
ScaleMaskAll
BG2
VZ4
TGT100
Alpha10.040000
Alpha20.060000
SF1.203084
NF1.000000
HZ4
Tau0.015000

Slide: Rayson_647-11_upf1-5_NMD_3hr_cordycepin_Rep1_ATH1

Add to Slide Selection

Get the data!

Summary BioSource 1 Information Extraction / Labelling Protocol for Source 1 Information Slide Information Hybridisation Analysis Information for "Affymetrix MAS 5.0 Scaling Protocol" normalised data.

Summary Information

This slide has had 1 extract(s) hybridised to it (1-colour).

  • Extract 1 ("Rayson_647-11_upf1-5_NMD_3hr_cordycepin_Rep1_ATH1") was made from "Arabidopsis thaliana" and labelled using "Biotin Labelled cRNA using Affymetrix Enzo Kit".

The slide used was "Affymetrix ATH1 Arabidopsis Genome Array".

Click on the options above for more information about this slide and its preparation.

Hybridisation Protocol Information

Name of Protocol: Affymetrix Antibody Amplification Protocol
Solution:
BufferEDTA, NaCl, Tween20 according to Affymetrix protocol
Water
Blocking Agent:
Herring Sperm DNA(Source: Promega)100pg/ml
Acetylated BSA(Source: Sigma-Aldrich)0.1mg/ml
Wash Procedure:
Step 1(Source: Affymetrix)10 cycles of 2 mixes/cycle with Non-Stringent buffer
Step 2(Source: Affymetrix)4 cycles of 15 mixes/cycle with Stringent buffer
Step 3(Source: Affymetrix)Stain the probe array for 10 minutes in Streptavaidin Phycoerythrin (SAPE) solution at 25 C
Step 4(Source: Affymetrix)10 cycles of 4 mixes/cycle with Non-Stringent buffer
Step 5(Source: Affymetrix)Stain the probe array for 10 minutes in antibody solution
Step 6(Source: Affymetrix)Stain the probe array for 10 minutes in SAPE solution at 25 C
Step 7(Source: Affymetrix)15 cycles of 4mixes/cycle with Non-Stringent buffer at 30 C
Step 8(Source: Affymetrix)Hold at 25 C
Quantity Used:
Amount15 microgram Fragmented cRNA
Time: 16 hours
Concentration: 50 pg/ml
Volume: 200 microlitres
Temperature: 45 C
Other Information:
Spike(Source: Affymetrix)Control Oligo B2 to 50pM for landing lights
Spike(Source: Affymetrix)20X Eukaryotic Hybridisation Controls (bioB (1.5pM, bioC 5pM, bioD 25pM, cre 100pM)
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

BioSource 1 Information

BioSource Name: Rayson_647-11_upf1-5_NMD_3hr_cordycepin_Rep1_ATH1
Organism: Arabidopsis thaliana
Alias: upf1-5
Stock Code: N9902
Genetic Background: Col-0
Age: 3 weeks
Growth Conditions:
StratificationImbibed in the dark at 4 °C for 3 days.
ProtocolPlants were grown in a Sanyo MLR-352 H with additive humidity under a typical short day cycle (8hr light:16 hr dark) The temperature was maintained at 21-23°C and the humidity was maintained at 65 % RH. Stratified seeds were sown onto SHL compost at a density of 4 seed per cell (24 cell tray). Trays were covered in a propagator lid, which was removed 7 days after sowing. Compost was kept moist throughout.
Soil ConstituentsSinclair Potting Growth Medium (www.william-sinclair.co.uk)
Plant Spacing1 per cell
waterCompost was kept moist at all times
Temperature22 °C average, 23 °C day, 21 °C night
Humidity65 % average, 65 % day, 65 % night
MediumNone
Lighting(Source: Cool white fluorescent manufactured by SANYO. Intensity: 175µEinsteins. Wavelength: White)
Developmental Stage:
Growth Stage(Source: Paradigm Genetics)
Tissue: Aerial tissues
in vivo Treatment: Time = 3 hours Aerial tissues were harvested into incubation buffer (IB: 1 mM Pipes, 1 mM Sodium citrate, 1 mM Potassium chloride, 15 mM Sucrose). After 30 minutes in IB, all except the time (T) = 0 samples were transferred to IB with cordycepin (150 µg ml-1). The T = 0 samples were transferred to fresh IB without cordycepin. All flasks (including the T = 0) were transferred to a bell jar and vacuum infiltrated. Immediately post-infiltration was classed as T =0.
Additional Organism Information:
Sample DescriptionSoil-grown plant grown under short-day conditions.
Other Information:
Timecourse start procedureThe time-course started immediately after the vacuum infiltration.

Protocols for BioSource 1

Extraction Protocol Information

Name of Protocol: Qiagen / Trizol
Method:
ProtocolQiagen RNAeasy kit protocol
Type: total RNA
Amplification: Amplified during labelling process - see labelling protocol
Other Information:
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

Labelling Protocol Information

Name of Protocol: Biotin Labelled cRNA using Affymetrix Enzo Kit
Amount Labelled: Unknown
Label Name: Biotin
Labelling Method: In-vitro transcription method
Other Information:
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

Slide Information

Date Hybridised:

Array Design Information

Slide Design Name: Affymetrix ATH1 Arabidopsis Genome Array
Array Type: Affymetrix Gene Chip
Number of Spots on the Array: 22810

Scanning Protocol Information

Name: Affymetrix MAS 5.0 Standard Scanning
Width of Image Produced (Pixels): 4733
Height of Image Produced (Pixels): 4733
X Resolution (microns): 3
Y Resolution (microns): 3
Scanner Name: Agilent 2500A GeneArray Scanner
Software Name: Affymetrix Micro Array Suite 5.0

Image Analysis Information

Scaling Factor:1.224611
Name: Affymetrix MAS 5.0 Standard Image Analysis
Software Name: Affymetrix Microarray Analysis Suite 5.0

Normalisation Information

Strategy Name: Affymetrix MAS 5.0 Scaling Protocol
Algorithm: The top 2% and bottom 2% of signal intensities are excluded, then the mean is calculated. The original signal values are scaled such that the mean is made equal to 100.
Control Elements:
none
Normalisation Protocol Parameters:
Spike_AFFX-r2-P1-cre_3-5-ratio1.235351
Spike_AFFX-r2-Ec-bioB_3-5-ratio0.998647
Spike_AFFX-r2-Bs-dap_5_signal64.830452
NoiseAvg:2.67,Std:0.09,Min:2.4,Max:2.9
Control DirectionAntisense
Spike_AFFX-r2-Ec-bioB_5_detectionP
Spike_AFFX-r2-Ec-bioB_3_detectionP
Spike_AFFX-r2-Bs-thr_5_signal34.334156
#P10842
Spike_AFFX-r2-Bs-phe_M_signal49.737827
Corner-Avg:10973,Count:32
Spike_AFFX-r2-P1-cre_3_detectionP
Spike_AFFX-r2-Ec-bioB_M_signal18.533289
Spike_AFFX-r2-Ec-bioB_3_signal26.195854
Spike_AFFX-r2-Bs-lys_M_signal42.413734
Spike_AFFX-r2-P1-cre_3_signal10053.690430
Spike_AFFX-r2-Bs-lys_3-5-ratio2.342619
Spike_AFFX-r2-Bs-dap_M_signal259.448761
Spike_AFFX-r2-Bs-thr_M_detectionP
Probe Pair Threshold1
Spike_AFFX-r2-Bs-thr_3-5-ratio6.819677
Spike_AFFX-r2-Ec-bioB_avg-signal23.653496
Spike_AFFX-r2-Bs-thr_avg-signal124.240662
Spike_AFFX-r2-Bs-thr_5_detectionP
Spike_AFFX-r2-Bs-dap_3_detectionP
Spike_AFFX-r2-Ec-bioD_5_signal2366.470947
Spike_AFFX-r2-Bs-phe_5_signal33.240116
Spike_AFFX-r2-P1-cre_5_detectionP
Spike_AFFX-r2-Ec-bioD_3_signal3108.045898
RawQ2.624514
Spike_AFFX-r2-Bs-lys_5_signal27.478043
Signal(A)8.289913
%A50.061378
Signal(All)172.352768
Corner+Avg:175,Count:32
Spike_AFFX-r2-Ec-bioD_5_detectionP
Spike_AFFX-r2-Ec-bioC_3_signal685.968140
Spike_AFFX-r2-Bs-lys_3_detectionP
Spike_AFFX-r2-Bs-phe_3_signal86.167000
Spike_AFFX-r2-Ec-bioD_avg-signal2737.258301
Spike_AFFX-r2-Bs-dap_5_detectionP
Spike_AFFX-r2-Bs-phe_5_detectionP
%P47.531784
Spike_AFFX-r2-Bs-lys_avg-signal44.754120
Spike_AFFX-r2-P1-cre_avg-signal9096.007813
Spike_AFFX-r2-Ec-bioB_M_detectionP
Spike_AFFX-r2-Bs-lys_M_detectionP
#Probe Sets Exceeding Probe Pair Threshold22810
%M2.406839
Spike_AFFX-r2-Bs-thr_3_signal234.147858
Spike_AFFX-r2-Ec-bioC_5_detectionP
Spike_AFFX-r2-Bs-phe_3_detectionP
Spike_AFFX-r2-Ec-bioC_3_detectionP
Spike_AFFX-r2-Bs-phe_avg-signal56.381649
Spike_AFFX-r2-Bs-dap_3_signal472.487701
Spike_AFFX-r2-Ec-bioD_3-5-ratio1.313367
Spike_AFFX-r2-Ec-bioD_3_detectionP
Spike_AFFX-r2-Ec-bioC_5_signal590.043823
#M549
Spike_AFFX-r2-Bs-lys_5_detectionP
Spike_AFFX-r2-Bs-lys_3_signal64.370575
Spike_AFFX-r2-Bs-dap_M_detectionP
Spike_AFFX-r2-Ec-bioC_3-5-ratio1.162572
Spike_AFFX-r2-Bs-thr_M_signal104.239960
Signal(P)352.500427
Spike_AFFX-r2-Bs-phe_3-5-ratio2.592259
Spike_AFFX-r2-Bs-thr_3_detectionP
Central-Avg:9700,Count:9
Spike_AFFX-r2-P1-cre_5_signal8138.325684
Spike_AFFX-r2-Bs-phe_M_detectionP
#A11419
Signal(M)27.129978
BackgroundAvg:66.02,Std:0.77,Min:64.5,Max:68.8
Spike_AFFX-r2-Ec-bioC_avg-signal638.005981
Spike_AFFX-r2-Bs-dap_avg-signal265.588959
Spike_AFFX-r2-Bs-dap_3-5-ratio7.288052
Spike_AFFX-r2-Ec-bioB_5_signal26.231344
ScaleMaskAll
BG2
VZ4
TGT100
Alpha10.040000
Alpha20.060000
SF1.224611
NF1.000000
HZ4
Tau0.015000

Slide: Rayson_647-10_upf1-5_NMD_1hr_cordycepin_Rep1_ATH1

Add to Slide Selection

Get the data!

Summary BioSource 1 Information Extraction / Labelling Protocol for Source 1 Information Slide Information Hybridisation Analysis Information for "Affymetrix MAS 5.0 Scaling Protocol" normalised data.

Summary Information

This slide has had 1 extract(s) hybridised to it (1-colour).

  • Extract 1 ("Rayson_647-10_upf1-5_NMD_1hr_cordycepin_Rep1_ATH1") was made from "Arabidopsis thaliana" and labelled using "Biotin Labelled cRNA using Affymetrix Enzo Kit".

The slide used was "Affymetrix ATH1 Arabidopsis Genome Array".

Click on the options above for more information about this slide and its preparation.

Hybridisation Protocol Information

Name of Protocol: Affymetrix Antibody Amplification Protocol
Solution:
BufferEDTA, NaCl, Tween20 according to Affymetrix protocol
Water
Blocking Agent:
Herring Sperm DNA(Source: Promega)100pg/ml
Acetylated BSA(Source: Sigma-Aldrich)0.1mg/ml
Wash Procedure:
Step 1(Source: Affymetrix)10 cycles of 2 mixes/cycle with Non-Stringent buffer
Step 2(Source: Affymetrix)4 cycles of 15 mixes/cycle with Stringent buffer
Step 3(Source: Affymetrix)Stain the probe array for 10 minutes in Streptavaidin Phycoerythrin (SAPE) solution at 25 C
Step 4(Source: Affymetrix)10 cycles of 4 mixes/cycle with Non-Stringent buffer
Step 5(Source: Affymetrix)Stain the probe array for 10 minutes in antibody solution
Step 6(Source: Affymetrix)Stain the probe array for 10 minutes in SAPE solution at 25 C
Step 7(Source: Affymetrix)15 cycles of 4mixes/cycle with Non-Stringent buffer at 30 C
Step 8(Source: Affymetrix)Hold at 25 C
Quantity Used:
Amount15 microgram Fragmented cRNA
Time: 16 hours
Concentration: 50 pg/ml
Volume: 200 microlitres
Temperature: 45 C
Other Information:
Spike(Source: Affymetrix)Control Oligo B2 to 50pM for landing lights
Spike(Source: Affymetrix)20X Eukaryotic Hybridisation Controls (bioB (1.5pM, bioC 5pM, bioD 25pM, cre 100pM)
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

BioSource 1 Information

BioSource Name: Rayson_647-10_upf1-5_NMD_1hr_cordycepin_Rep1_ATH1
Organism: Arabidopsis thaliana
Alias: upf1-5
Stock Code: N9902
Genetic Background: Col-0
Age: 3 weeks
Growth Conditions:
StratificationImbibed in the dark at 4 °C for 3 days.
ProtocolPlants were grown in a Sanyo MLR-352 H with additive humidity under a typical short day cycle (8hr light:16 hr dark) The temperature was maintained at 21-23°C and the humidity was maintained at 65 % RH. Stratified seeds were sown onto SHL compost at a density of 4 seed per cell (24 cell tray). Trays were covered in a propagator lid, which was removed 7 days after sowing. Compost was kept moist throughout.
Soil ConstituentsSinclair Potting Growth Medium (www.william-sinclair.co.uk)
Plant Spacing1 per cell
waterCompost was kept moist at all times
Temperature22 °C average, 23 °C day, 21 °C night
Humidity65 % average, 65 % day, 65 % night
MediumNone
Lighting(Source: Cool white fluorescent manufactured by SANYO. Intensity: 175µEinsteins. Wavelength: White)
Developmental Stage:
Growth Stage(Source: Paradigm Genetics)
Tissue: Aerial tissues
in vivo Treatment: Time = 1 hour Aerial tissues were harvested into incubation buffer (IB: 1 mM Pipes, 1 mM Sodium citrate, 1 mM Potassium chloride, 15 mM Sucrose). After 30 minutes in IB, all except the time (T) = 0 samples were transferred to IB with cordycepin (150 µg ml-1). The T = 0 samples were transferred to fresh IB without cordycepin. All flasks (including the T = 0) were transferred to a bell jar and vacuum infiltrated. Immediately post-infiltration was classed as T =0.
Additional Organism Information:
Sample DescriptionSoil-grown plant grown under short-day conditions.
Other Information:
Timecourse start procedureThe time-course started immediately after the vacuum infiltration.

Protocols for BioSource 1

Extraction Protocol Information

Name of Protocol: Qiagen / Trizol
Method:
ProtocolQiagen RNAeasy kit protocol
Type: total RNA
Amplification: Amplified during labelling process - see labelling protocol
Other Information:
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

Labelling Protocol Information

Name of Protocol: Biotin Labelled cRNA using Affymetrix Enzo Kit
Amount Labelled: Unknown
Label Name: Biotin
Labelling Method: In-vitro transcription method
Other Information:
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

Slide Information

Date Hybridised:

Array Design Information

Slide Design Name: Affymetrix ATH1 Arabidopsis Genome Array
Array Type: Affymetrix Gene Chip
Number of Spots on the Array: 22810

Scanning Protocol Information

Name: Affymetrix MAS 5.0 Standard Scanning
Width of Image Produced (Pixels): 4733
Height of Image Produced (Pixels): 4733
X Resolution (microns): 3
Y Resolution (microns): 3
Scanner Name: Agilent 2500A GeneArray Scanner
Software Name: Affymetrix Micro Array Suite 5.0

Image Analysis Information

Scaling Factor:1.10675
Name: Affymetrix MAS 5.0 Standard Image Analysis
Software Name: Affymetrix Microarray Analysis Suite 5.0

Normalisation Information

Strategy Name: Affymetrix MAS 5.0 Scaling Protocol
Algorithm: The top 2% and bottom 2% of signal intensities are excluded, then the mean is calculated. The original signal values are scaled such that the mean is made equal to 100.
Control Elements:
none
Normalisation Protocol Parameters:
Spike_AFFX-r2-P1-cre_3-5-ratio1.204247
Spike_AFFX-r2-Ec-bioB_3-5-ratio2.323662
Spike_AFFX-r2-Bs-dap_5_signal46.152069
NoiseAvg:2.76,Std:0.05,Min:2.6,Max:2.9
Control DirectionAntisense
Spike_AFFX-r2-Ec-bioB_5_detectionA
Spike_AFFX-r2-Ec-bioB_3_detectionP
Spike_AFFX-r2-Bs-thr_5_signal12.660348
#P12260
Spike_AFFX-r2-Bs-phe_M_signal27.773920
Corner-Avg:10708,Count:32
Spike_AFFX-r2-P1-cre_3_detectionP
Spike_AFFX-r2-Ec-bioB_M_signal27.977093
Spike_AFFX-r2-Ec-bioB_3_signal33.010414
Spike_AFFX-r2-Bs-lys_M_signal15.801888
Spike_AFFX-r2-P1-cre_3_signal8798.389648
Spike_AFFX-r2-Bs-lys_3-5-ratio3.062706
Spike_AFFX-r2-Bs-dap_M_signal148.549683
Spike_AFFX-r2-Bs-thr_M_detectionP
Probe Pair Threshold1
Spike_AFFX-r2-Bs-thr_3-5-ratio9.881851
Spike_AFFX-r2-Ec-bioB_avg-signal25.064569
Spike_AFFX-r2-Bs-thr_avg-signal67.030861
Spike_AFFX-r2-Bs-thr_5_detectionA
Spike_AFFX-r2-Bs-dap_3_detectionP
Spike_AFFX-r2-Ec-bioD_5_signal2198.581299
Spike_AFFX-r2-Bs-phe_5_signal17.877298
Spike_AFFX-r2-P1-cre_5_detectionP
Spike_AFFX-r2-Ec-bioD_3_signal2598.280029
RawQ2.722867
Spike_AFFX-r2-Bs-lys_5_signal15.183534
Signal(A)7.013395
%A44.072777
Signal(All)158.762894
Corner+Avg:192,Count:32
Spike_AFFX-r2-Ec-bioD_5_detectionP
Spike_AFFX-r2-Ec-bioC_3_signal542.471313
Spike_AFFX-r2-Bs-lys_3_detectionP
Spike_AFFX-r2-Bs-phe_3_signal64.417236
Spike_AFFX-r2-Ec-bioD_avg-signal2398.430664
Spike_AFFX-r2-Bs-dap_5_detectionP
Spike_AFFX-r2-Bs-phe_5_detectionP
%P53.748356
Spike_AFFX-r2-Bs-lys_avg-signal25.829374
Spike_AFFX-r2-P1-cre_avg-signal8052.261719
Spike_AFFX-r2-Ec-bioB_M_detectionP
Spike_AFFX-r2-Bs-lys_M_detectionP
#Probe Sets Exceeding Probe Pair Threshold22810
%M2.178869
Spike_AFFX-r2-Bs-thr_3_signal125.107674
Spike_AFFX-r2-Ec-bioC_5_detectionP
Spike_AFFX-r2-Bs-phe_3_detectionP
Spike_AFFX-r2-Ec-bioC_3_detectionP
Spike_AFFX-r2-Bs-phe_avg-signal36.689484
Spike_AFFX-r2-Bs-dap_3_signal280.245667
Spike_AFFX-r2-Ec-bioD_3-5-ratio1.181798
Spike_AFFX-r2-Ec-bioD_3_detectionP
Spike_AFFX-r2-Ec-bioC_5_signal584.345581
#M497
Spike_AFFX-r2-Bs-lys_5_detectionP
Spike_AFFX-r2-Bs-lys_3_signal46.502701
Spike_AFFX-r2-Bs-dap_M_detectionP
Spike_AFFX-r2-Ec-bioC_3-5-ratio0.928340
Spike_AFFX-r2-Bs-thr_M_signal63.324543
Signal(P)288.584930
Spike_AFFX-r2-Bs-phe_3-5-ratio3.603298
Spike_AFFX-r2-Bs-thr_3_detectionP
Central-Avg:8130,Count:9
Spike_AFFX-r2-P1-cre_5_signal7306.134277
Spike_AFFX-r2-Bs-phe_M_detectionM
#A10053
Signal(M)25.804132
BackgroundAvg:69.03,Std:0.63,Min:68.3,Max:71.7
Spike_AFFX-r2-Ec-bioC_avg-signal563.408447
Spike_AFFX-r2-Bs-dap_avg-signal158.315811
Spike_AFFX-r2-Bs-dap_3-5-ratio6.072223
Spike_AFFX-r2-Ec-bioB_5_signal14.206201
ScaleMaskAll
BG2
VZ4
TGT100
Alpha10.040000
Alpha20.060000
SF1.106750
NF1.000000
HZ4
Tau0.015000

Slide: Rayson_647-15_UPF2(6-11E5)_NMD_3hr _cordycepin_Rep1_ATH1

Add to Slide Selection

Get the data!

Summary BioSource 1 Information Extraction / Labelling Protocol for Source 1 Information Slide Information Hybridisation Analysis Information for "Affymetrix MAS 5.0 Scaling Protocol" normalised data.

Summary Information

This slide has had 1 extract(s) hybridised to it (1-colour).

  • Extract 1 ("Rayson_647-15_UPF2(6-11E5)_NMD_3hr _cordycepin_Rep1_ATH1") was made from "Arabidopsis thaliana" and labelled using "Biotin Labelled cRNA using Affymetrix Enzo Kit".

The slide used was "Affymetrix ATH1 Arabidopsis Genome Array".

Click on the options above for more information about this slide and its preparation.

Hybridisation Protocol Information

Name of Protocol: Affymetrix Antibody Amplification Protocol
Solution:
BufferEDTA, NaCl, Tween20 according to Affymetrix protocol
Water
Blocking Agent:
Herring Sperm DNA(Source: Promega)100pg/ml
Acetylated BSA(Source: Sigma-Aldrich)0.1mg/ml
Wash Procedure:
Step 1(Source: Affymetrix)10 cycles of 2 mixes/cycle with Non-Stringent buffer
Step 2(Source: Affymetrix)4 cycles of 15 mixes/cycle with Stringent buffer
Step 3(Source: Affymetrix)Stain the probe array for 10 minutes in Streptavaidin Phycoerythrin (SAPE) solution at 25 C
Step 4(Source: Affymetrix)10 cycles of 4 mixes/cycle with Non-Stringent buffer
Step 5(Source: Affymetrix)Stain the probe array for 10 minutes in antibody solution
Step 6(Source: Affymetrix)Stain the probe array for 10 minutes in SAPE solution at 25 C
Step 7(Source: Affymetrix)15 cycles of 4mixes/cycle with Non-Stringent buffer at 30 C
Step 8(Source: Affymetrix)Hold at 25 C
Quantity Used:
Amount15 microgram Fragmented cRNA
Time: 16 hours
Concentration: 50 pg/ml
Volume: 200 microlitres
Temperature: 45 C
Other Information:
Spike(Source: Affymetrix)Control Oligo B2 to 50pM for landing lights
Spike(Source: Affymetrix)20X Eukaryotic Hybridisation Controls (bioB (1.5pM, bioC 5pM, bioD 25pM, cre 100pM)
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

BioSource 1 Information

BioSource Name: Rayson_647-15_UPF2(6-11E5)_NMD_3hr _cordycepin_Rep1_ATH1
Organism: Arabidopsis thaliana
Alias: 6-11E5
Stock Code:
Age: 3 weeks
Growth Conditions:
StratificationImbibed in the dark at 4 °C for 3 days.
ProtocolPlants were grown in a Sanyo MLR-352 H with additive humidity under a typical short day cycle (8hr light:16 hr dark) The temperature was maintained at 21-23°C and the humidity was maintained at 65 % RH. Stratified seeds were sown onto SHL compost at a density of 4 seed per cell (24 cell tray). Trays were covered in a propagator lid, which was removed 7 days after sowing. Compost was kept moist throughout.
Soil ConstituentsSinclair Potting Growth Medium (www.william-sinclair.co.uk)
Plant Spacing1 per cell
waterCompost was kept moist at all times
Temperature22 °C average, 23 °C day, 21 °C night
Humidity65 % average, 65 % day, 65 % night
MediumNone
Lighting(Source: Cool white fluorescent manufactured by SANYO. Intensity: 175µEinsteins. Wavelength: White)
Developmental Stage:
Growth Stage(Source: Paradigm Genetics)
Genetic Variation: induced mutation mutation in At2g39260
Tissue: Aerial tissues
in vivo Treatment: Time = 3 hours Aerial tissues were harvested into incubation buffer (IB: 1 mM Pipes, 1 mM Sodium citrate, 1 mM Potassium chloride, 15 mM Sucrose). After 30 minutes in IB, all except the time (T) = 0 samples were transferred to IB with cordycepin (150 µg ml-1). The T = 0 samples were transferred to fresh IB without cordycepin. All flasks (including the T = 0) were transferred to a bell jar and vacuum infiltrated. Immediately post-infiltration was classed as T =0.
Additional Organism Information:
Sample DescriptionSoil-grown plant grown under short-day conditions.
Other Information:
Timecourse start procedureThe time-course started immediately after the vacuum infiltration.

Protocols for BioSource 1

Extraction Protocol Information

Name of Protocol: Qiagen / Trizol
Method:
ProtocolQiagen RNAeasy kit protocol
Type: total RNA
Amplification: Amplified during labelling process - see labelling protocol
Other Information:
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

Labelling Protocol Information

Name of Protocol: Biotin Labelled cRNA using Affymetrix Enzo Kit
Amount Labelled: Unknown
Label Name: Biotin
Labelling Method: In-vitro transcription method
Other Information:
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

Slide Information

Date Hybridised:

Array Design Information

Slide Design Name: Affymetrix ATH1 Arabidopsis Genome Array
Array Type: Affymetrix Gene Chip
Number of Spots on the Array: 22810

Scanning Protocol Information

Name: Affymetrix MAS 5.0 Standard Scanning
Width of Image Produced (Pixels): 4733
Height of Image Produced (Pixels): 4733
X Resolution (microns): 3
Y Resolution (microns): 3
Scanner Name: Agilent 2500A GeneArray Scanner
Software Name: Affymetrix Micro Array Suite 5.0

Image Analysis Information

Scaling Factor:0.656874
Name: Affymetrix MAS 5.0 Standard Image Analysis
Software Name: Affymetrix Microarray Analysis Suite 5.0

Normalisation Information

Strategy Name: Affymetrix MAS 5.0 Scaling Protocol
Algorithm: The top 2% and bottom 2% of signal intensities are excluded, then the mean is calculated. The original signal values are scaled such that the mean is made equal to 100.
Control Elements:
none
Normalisation Protocol Parameters:
Spike_AFFX-r2-P1-cre_3-5-ratio1.344727
Spike_AFFX-r2-Ec-bioB_3-5-ratio1.031250
Spike_AFFX-r2-Bs-dap_5_signal26.047722
NoiseAvg:3.41,Std:0.10,Min:3.1,Max:3.8
Control DirectionAntisense
Spike_AFFX-r2-Ec-bioB_5_detectionA
Spike_AFFX-r2-Ec-bioB_3_detectionA
Spike_AFFX-r2-Bs-thr_5_signal11.998852
#P12779
Spike_AFFX-r2-Bs-phe_M_signal22.495623
Corner-Avg:11054,Count:32
Spike_AFFX-r2-P1-cre_3_detectionP
Spike_AFFX-r2-Ec-bioB_M_signal10.231617
Spike_AFFX-r2-Ec-bioB_3_signal10.449628
Spike_AFFX-r2-Bs-lys_M_signal13.628344
Spike_AFFX-r2-P1-cre_3_signal4553.129395
Spike_AFFX-r2-Bs-lys_3-5-ratio1.908774
Spike_AFFX-r2-Bs-dap_M_signal137.534485
Spike_AFFX-r2-Bs-thr_M_detectionP
Probe Pair Threshold1
Spike_AFFX-r2-Bs-thr_3-5-ratio8.163610
Spike_AFFX-r2-Ec-bioB_avg-signal10.271405
Spike_AFFX-r2-Bs-thr_avg-signal47.349808
Spike_AFFX-r2-Bs-thr_5_detectionA
Spike_AFFX-r2-Bs-dap_3_detectionP
Spike_AFFX-r2-Ec-bioD_5_signal702.139343
Spike_AFFX-r2-Bs-phe_5_signal17.138174
Spike_AFFX-r2-P1-cre_5_detectionP
Spike_AFFX-r2-Ec-bioD_3_signal1311.920166
RawQ3.028281
Spike_AFFX-r2-Bs-lys_5_signal15.811852
Signal(A)5.242609
%A41.994740
Signal(All)148.630829
Corner+Avg:218,Count:32
Spike_AFFX-r2-Ec-bioD_5_detectionP
Spike_AFFX-r2-Ec-bioC_3_signal233.618530
Spike_AFFX-r2-Bs-lys_3_detectionP
Spike_AFFX-r2-Bs-phe_3_signal46.787178
Spike_AFFX-r2-Ec-bioD_avg-signal1007.029785
Spike_AFFX-r2-Bs-dap_5_detectionP
Spike_AFFX-r2-Bs-phe_5_detectionP
%P56.023674
Spike_AFFX-r2-Bs-lys_avg-signal19.873817
Spike_AFFX-r2-P1-cre_avg-signal3969.521973
Spike_AFFX-r2-Ec-bioB_M_detectionP
Spike_AFFX-r2-Bs-lys_M_detectionP
#Probe Sets Exceeding Probe Pair Threshold22810
%M1.981587
Spike_AFFX-r2-Bs-thr_3_signal97.953949
Spike_AFFX-r2-Ec-bioC_5_detectionP
Spike_AFFX-r2-Bs-phe_3_detectionP
Spike_AFFX-r2-Ec-bioC_3_detectionP
Spike_AFFX-r2-Bs-phe_avg-signal28.806992
Spike_AFFX-r2-Bs-dap_3_signal201.975510
Spike_AFFX-r2-Ec-bioD_3-5-ratio1.868461
Spike_AFFX-r2-Ec-bioD_3_detectionP
Spike_AFFX-r2-Ec-bioC_5_signal221.348221
#M452
Spike_AFFX-r2-Bs-lys_5_detectionP
Spike_AFFX-r2-Bs-lys_3_signal30.181259
Spike_AFFX-r2-Bs-dap_M_detectionP
Spike_AFFX-r2-Ec-bioC_3-5-ratio1.055434
Spike_AFFX-r2-Bs-thr_M_signal32.096622
Signal(P)260.716431
Spike_AFFX-r2-Bs-phe_3-5-ratio2.729998
Spike_AFFX-r2-Bs-thr_3_detectionP
Central-Avg:9043,Count:9
Spike_AFFX-r2-P1-cre_5_signal3385.914551
Spike_AFFX-r2-Bs-phe_M_detectionP
#A9579
Signal(M)18.484840
BackgroundAvg:82.58,Std:0.93,Min:80.2,Max:85.6
Spike_AFFX-r2-Ec-bioC_avg-signal227.483368
Spike_AFFX-r2-Bs-dap_avg-signal121.852577
Spike_AFFX-r2-Bs-dap_3-5-ratio7.754056
Spike_AFFX-r2-Ec-bioB_5_signal10.132971
ScaleMaskAll
BG2
VZ4
TGT100
Alpha10.040000
Alpha20.060000
SF0.656874
NF1.000000
HZ4
Tau0.015000

Slide: Rayson_647-14_UPF2(6-11E5)_NMD_1hr _cordycepin_Rep1_ATH1

Add to Slide Selection

Get the data!

Summary BioSource 1 Information Extraction / Labelling Protocol for Source 1 Information Slide Information Hybridisation Analysis Information for "Affymetrix MAS 5.0 Scaling Protocol" normalised data.

Summary Information

This slide has had 1 extract(s) hybridised to it (1-colour).

  • Extract 1 ("Rayson_647-14_UPF2(6-11E5)_NMD_1hr _cordycepin_Rep1_ATH1") was made from "Arabidopsis thaliana" and labelled using "Biotin Labelled cRNA using Affymetrix Enzo Kit".

The slide used was "Affymetrix ATH1 Arabidopsis Genome Array".

Click on the options above for more information about this slide and its preparation.

Hybridisation Protocol Information

Name of Protocol: Affymetrix Antibody Amplification Protocol
Solution:
BufferEDTA, NaCl, Tween20 according to Affymetrix protocol
Water
Blocking Agent:
Herring Sperm DNA(Source: Promega)100pg/ml
Acetylated BSA(Source: Sigma-Aldrich)0.1mg/ml
Wash Procedure:
Step 1(Source: Affymetrix)10 cycles of 2 mixes/cycle with Non-Stringent buffer
Step 2(Source: Affymetrix)4 cycles of 15 mixes/cycle with Stringent buffer
Step 3(Source: Affymetrix)Stain the probe array for 10 minutes in Streptavaidin Phycoerythrin (SAPE) solution at 25 C
Step 4(Source: Affymetrix)10 cycles of 4 mixes/cycle with Non-Stringent buffer
Step 5(Source: Affymetrix)Stain the probe array for 10 minutes in antibody solution
Step 6(Source: Affymetrix)Stain the probe array for 10 minutes in SAPE solution at 25 C
Step 7(Source: Affymetrix)15 cycles of 4mixes/cycle with Non-Stringent buffer at 30 C
Step 8(Source: Affymetrix)Hold at 25 C
Quantity Used:
Amount15 microgram Fragmented cRNA
Time: 16 hours
Concentration: 50 pg/ml
Volume: 200 microlitres
Temperature: 45 C
Other Information:
Spike(Source: Affymetrix)Control Oligo B2 to 50pM for landing lights
Spike(Source: Affymetrix)20X Eukaryotic Hybridisation Controls (bioB (1.5pM, bioC 5pM, bioD 25pM, cre 100pM)
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

BioSource 1 Information

BioSource Name: Rayson_647-14_UPF2(6-11E5)_NMD_1hr _cordycepin_Rep1_ATH1
Organism: Arabidopsis thaliana
Alias: 6-11E5
Stock Code:
Age: 3 weeks
Growth Conditions:
StratificationImbibed in the dark at 4 °C for 3 days.
ProtocolPlants were grown in a Sanyo MLR-352 H with additive humidity under a typical short day cycle (8hr light:16 hr dark) The temperature was maintained at 21-23°C and the humidity was maintained at 65 % RH. Stratified seeds were sown onto SHL compost at a density of 4 seed per cell (24 cell tray). Trays were covered in a propagator lid, which was removed 7 days after sowing. Compost was kept moist throughout.
Soil ConstituentsSinclair Potting Growth Medium (www.william-sinclair.co.uk)
Plant Spacing1 per cell
waterCompost was kept moist at all times
Temperature22 °C average, 23 °C day, 21 °C night
Humidity65 % average, 65 % day, 65 % night
MediumNone
Lighting(Source: Cool white fluorescent manufactured by SANYO. Intensity: 175µEinsteins. Wavelength: White)
Developmental Stage:
Growth Stage(Source: Paradigm Genetics)
Genetic Variation: induced mutation mutation in At2g39260
Tissue: Aerial tissues
in vivo Treatment: Time = 1 hour Aerial tissues were harvested into incubation buffer (IB: 1 mM Pipes, 1 mM Sodium citrate, 1 mM Potassium chloride, 15 mM Sucrose). After 30 minutes in IB, all except the time (T) = 0 samples were transferred to IB with cordycepin (150 µg ml-1). The T = 0 samples were transferred to fresh IB without cordycepin. All flasks (including the T = 0) were transferred to a bell jar and vacuum infiltrated. Immediately post-infiltration was classed as T =0.
Additional Organism Information:
Sample DescriptionSoil-grown plant grown under short-day conditions.
Other Information:
Timecourse start procedureThe time-course started immediately after the vacuum infiltration.

Protocols for BioSource 1

Extraction Protocol Information

Name of Protocol: Qiagen / Trizol
Method:
ProtocolQiagen RNAeasy kit protocol
Type: total RNA
Amplification: Amplified during labelling process - see labelling protocol
Other Information:
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

Labelling Protocol Information

Name of Protocol: Biotin Labelled cRNA using Affymetrix Enzo Kit
Amount Labelled: Unknown
Label Name: Biotin
Labelling Method: In-vitro transcription method
Other Information:
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

Slide Information

Date Hybridised:

Array Design Information

Slide Design Name: Affymetrix ATH1 Arabidopsis Genome Array
Array Type: Affymetrix Gene Chip
Number of Spots on the Array: 22810

Scanning Protocol Information

Name: Affymetrix MAS 5.0 Standard Scanning
Width of Image Produced (Pixels): 4733
Height of Image Produced (Pixels): 4733
X Resolution (microns): 3
Y Resolution (microns): 3
Scanner Name: Agilent 2500A GeneArray Scanner
Software Name: Affymetrix Micro Array Suite 5.0

Image Analysis Information

Scaling Factor:0.485274
Name: Affymetrix MAS 5.0 Standard Image Analysis
Software Name: Affymetrix Microarray Analysis Suite 5.0

Normalisation Information

Strategy Name: Affymetrix MAS 5.0 Scaling Protocol
Algorithm: The top 2% and bottom 2% of signal intensities are excluded, then the mean is calculated. The original signal values are scaled such that the mean is made equal to 100.
Control Elements:
none
Normalisation Protocol Parameters:
Spike_AFFX-r2-P1-cre_3-5-ratio1.219548
Spike_AFFX-r2-Ec-bioB_3-5-ratio3.283607
Spike_AFFX-r2-Bs-dap_5_signal31.983187
NoiseAvg:3.47,Std:0.17,Min:3.0,Max:4.1
Control DirectionAntisense
Spike_AFFX-r2-Ec-bioB_5_detectionA
Spike_AFFX-r2-Ec-bioB_3_detectionP
Spike_AFFX-r2-Bs-thr_5_signal15.758619
#P13536
Spike_AFFX-r2-Bs-phe_M_signal30.971397
Corner-Avg:11317,Count:32
Spike_AFFX-r2-P1-cre_3_detectionP
Spike_AFFX-r2-Ec-bioB_M_signal1.380003
Spike_AFFX-r2-Ec-bioB_3_signal14.634616
Spike_AFFX-r2-Bs-lys_M_signal17.869467
Spike_AFFX-r2-P1-cre_3_signal3411.189209
Spike_AFFX-r2-Bs-lys_3-5-ratio1.845810
Spike_AFFX-r2-Bs-dap_M_signal158.042786
Spike_AFFX-r2-Bs-thr_M_detectionP
Probe Pair Threshold1
Spike_AFFX-r2-Bs-thr_3-5-ratio7.376622
Spike_AFFX-r2-Ec-bioB_avg-signal6.823830
Spike_AFFX-r2-Bs-thr_avg-signal62.525990
Spike_AFFX-r2-Bs-thr_5_detectionP
Spike_AFFX-r2-Bs-dap_3_detectionP
Spike_AFFX-r2-Ec-bioD_5_signal639.675476
Spike_AFFX-r2-Bs-phe_5_signal23.193953
Spike_AFFX-r2-P1-cre_5_detectionP
Spike_AFFX-r2-Ec-bioD_3_signal1034.489014
RawQ3.103346
Spike_AFFX-r2-Bs-lys_5_signal21.869366
Signal(A)4.278897
%A38.882069
Signal(All)142.488235
Corner+Avg:196,Count:32
Spike_AFFX-r2-Ec-bioD_5_detectionP
Spike_AFFX-r2-Ec-bioC_3_signal174.229614
Spike_AFFX-r2-Bs-lys_3_detectionP
Spike_AFFX-r2-Bs-phe_3_signal53.081852
Spike_AFFX-r2-Ec-bioD_avg-signal837.082275
Spike_AFFX-r2-Bs-dap_5_detectionP
Spike_AFFX-r2-Bs-phe_5_detectionP
%P59.342392
Spike_AFFX-r2-Bs-lys_avg-signal26.701838
Spike_AFFX-r2-P1-cre_avg-signal3104.141113
Spike_AFFX-r2-Ec-bioB_M_detectionA
Spike_AFFX-r2-Bs-lys_M_detectionP
#Probe Sets Exceeding Probe Pair Threshold22810
%M1.775537
Spike_AFFX-r2-Bs-thr_3_signal116.245377
Spike_AFFX-r2-Ec-bioC_5_detectionP
Spike_AFFX-r2-Bs-phe_3_detectionP
Spike_AFFX-r2-Ec-bioC_3_detectionP
Spike_AFFX-r2-Bs-phe_avg-signal35.749069
Spike_AFFX-r2-Bs-dap_3_signal254.633087
Spike_AFFX-r2-Ec-bioD_3-5-ratio1.617209
Spike_AFFX-r2-Ec-bioD_3_detectionP
Spike_AFFX-r2-Ec-bioC_5_signal170.116058
#M405
Spike_AFFX-r2-Bs-lys_5_detectionP
Spike_AFFX-r2-Bs-lys_3_signal40.366688
Spike_AFFX-r2-Bs-dap_M_detectionP
Spike_AFFX-r2-Ec-bioC_3-5-ratio1.024181
Spike_AFFX-r2-Bs-thr_M_signal55.573975
Signal(P)236.821487
Spike_AFFX-r2-Bs-phe_3-5-ratio2.288607
Spike_AFFX-r2-Bs-thr_3_detectionP
Central-Avg:9500,Count:9
Spike_AFFX-r2-P1-cre_5_signal2797.093262
Spike_AFFX-r2-Bs-phe_M_detectionP
#A8869
Signal(M)16.274622
BackgroundAvg:81.10,Std:1.28,Min:77.8,Max:84.4
Spike_AFFX-r2-Ec-bioC_avg-signal172.172836
Spike_AFFX-r2-Bs-dap_avg-signal148.219681
Spike_AFFX-r2-Bs-dap_3-5-ratio7.961467
Spike_AFFX-r2-Ec-bioB_5_signal4.456872
ScaleMaskAll
BG2
VZ4
TGT100
Alpha10.040000
Alpha20.060000
SF0.485274
NF1.000000
HZ4
Tau0.015000

Slide: Rayson_647-13_UPF2(6-11E5)_NMD_0hr_cordycepin_Rep1_ATH1

Add to Slide Selection

Get the data!

Summary BioSource 1 Information Extraction / Labelling Protocol for Source 1 Information Slide Information Hybridisation Analysis Information for "Affymetrix MAS 5.0 Scaling Protocol" normalised data.

Summary Information

This slide has had 1 extract(s) hybridised to it (1-colour).

  • Extract 1 ("Rayson_647-13_UPF2(6-11E5)_NMD_0hr_cordycepin_Rep1_ATH1") was made from "Arabidopsis thaliana" and labelled using "Biotin Labelled cRNA using Affymetrix Enzo Kit".

The slide used was "Affymetrix ATH1 Arabidopsis Genome Array".

Click on the options above for more information about this slide and its preparation.

Hybridisation Protocol Information

Name of Protocol: Affymetrix Antibody Amplification Protocol
Solution:
BufferEDTA, NaCl, Tween20 according to Affymetrix protocol
Water
Blocking Agent:
Herring Sperm DNA(Source: Promega)100pg/ml
Acetylated BSA(Source: Sigma-Aldrich)0.1mg/ml
Wash Procedure:
Step 1(Source: Affymetrix)10 cycles of 2 mixes/cycle with Non-Stringent buffer
Step 2(Source: Affymetrix)4 cycles of 15 mixes/cycle with Stringent buffer
Step 3(Source: Affymetrix)Stain the probe array for 10 minutes in Streptavaidin Phycoerythrin (SAPE) solution at 25 C
Step 4(Source: Affymetrix)10 cycles of 4 mixes/cycle with Non-Stringent buffer
Step 5(Source: Affymetrix)Stain the probe array for 10 minutes in antibody solution
Step 6(Source: Affymetrix)Stain the probe array for 10 minutes in SAPE solution at 25 C
Step 7(Source: Affymetrix)15 cycles of 4mixes/cycle with Non-Stringent buffer at 30 C
Step 8(Source: Affymetrix)Hold at 25 C
Quantity Used:
Amount15 microgram Fragmented cRNA
Time: 16 hours
Concentration: 50 pg/ml
Volume: 200 microlitres
Temperature: 45 C
Other Information:
Spike(Source: Affymetrix)Control Oligo B2 to 50pM for landing lights
Spike(Source: Affymetrix)20X Eukaryotic Hybridisation Controls (bioB (1.5pM, bioC 5pM, bioD 25pM, cre 100pM)
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

BioSource 1 Information

BioSource Name: Rayson_647-13_UPF2(6-11E5)_NMD_0hr_cordycepin_Rep1_ATH1
Organism: Arabidopsis thaliana
Alias: 6-11E5
Stock Code:
Age: 3 weeks
Growth Conditions:
StratificationImbibed in the dark at 4 °C for 3 days.
ProtocolPlants were grown in a Sanyo MLR-352 H with additive humidity under a typical short day cycle (8hr light:16 hr dark) The temperature was maintained at 21-23°C and the humidity was maintained at 65 % RH. Stratified seeds were sown onto SHL compost at a density of 4 seed per cell (24 cell tray). Trays were covered in a propagator lid, which was removed 7 days after sowing. Compost was kept moist throughout.
Soil ConstituentsSinclair Potting Growth Medium (www.william-sinclair.co.uk)
Plant Spacing1 per cell
waterCompost was kept moist at all times
Temperature22 °C average, 23 °C day, 21 °C night
Humidity65 % average, 65 % day, 65 % night
MediumNone
Lighting(Source: Cool white fluorescent manufactured by SANYO. Intensity: 175µEinsteins. Wavelength: White)
Developmental Stage:
Growth Stage(Source: Paradigm Genetics)
Genetic Variation: induced mutation mutation in At2g39260
Tissue: Aerial tissues
in vivo Treatment: Time = 0 Aerial tissues were harvested into incubation buffer (IB: 1 mM Pipes, 1 mM Sodium citrate, 1 mM Potassium chloride, 15 mM Sucrose). After 30 minutes in IB, all except the time (T) = 0 samples were transferred to IB with cordycepin (150 µg ml-1). The T = 0 samples were transferred to fresh IB without cordycepin. All flasks (including the T = 0) were transferred to a bell jar and vacuum infiltrated. Immediately post-infiltration was classed as T =0.
Additional Organism Information:
Sample DescriptionSoil-grown plant grown under short-day conditions.
Other Information:
Timecourse start procedureThe time-course started immediately after the vacuum infiltration.

Protocols for BioSource 1

Extraction Protocol Information

Name of Protocol: Qiagen / Trizol
Method:
ProtocolQiagen RNAeasy kit protocol
Type: total RNA
Amplification: Amplified during labelling process - see labelling protocol
Other Information:
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

Labelling Protocol Information

Name of Protocol: Biotin Labelled cRNA using Affymetrix Enzo Kit
Amount Labelled: Unknown
Label Name: Biotin
Labelling Method: In-vitro transcription method
Other Information:
ProtocolSourceAffymetrix GeneChip Technical Analysis Manual

Slide Information

Date Hybridised:

Array Design Information

Slide Design Name: Affymetrix ATH1 Arabidopsis Genome Array
Array Type: Affymetrix Gene Chip
Number of Spots on the Array: 22810

Scanning Protocol Information

Name: Affymetrix MAS 5.0 Standard Scanning
Width of Image Produced (Pixels): 4733
Height of Image Produced (Pixels): 4733
X Resolution (microns): 3
Y Resolution (microns): 3
Scanner Name: Agilent 2500A GeneArray Scanner
Software Name: Affymetrix Micro Array Suite 5.0

Image Analysis Information

Scaling Factor:0.359765
Name: Affymetrix MAS 5.0 Standard Image Analysis
Software Name: Affymetrix Microarray Analysis Suite 5.0

Normalisation Information

Strategy Name: Affymetrix MAS 5.0 Scaling Protocol
Algorithm: The top 2% and bottom 2% of signal intensities are excluded, then the mean is calculated. The original signal values are scaled such that the mean is made equal to 100.
Control Elements:
none
Normalisation Protocol Parameters:
Spike_AFFX-r2-P1-cre_3-5-ratio1.245371
Spike_AFFX-r2-Ec-bioB_3-5-ratio1.716131
Spike_AFFX-r2-Bs-dap_5_signal17.938309
NoiseAvg:4.22,Std:0.10,Min:3.9,Max:4.5
Control DirectionAntisense
Spike_AFFX-r2-Ec-bioB_5_detectionA
Spike_AFFX-r2-Ec-bioB_3_detectionA
Spike_AFFX-r2-Bs-thr_5_signal7.757352
#P13932
Spike_AFFX-r2-Bs-phe_M_signal15.116912
Corner-Avg:11664,Count:32
Spike_AFFX-r2-P1-cre_3_detectionP
Spike_AFFX-r2-Ec-bioB_M_signal8.405900
Spike_AFFX-r2-Ec-bioB_3_signal7.135137
Spike_AFFX-r2-Bs-lys_M_signal10.767626
Spike_AFFX-r2-P1-cre_3_signal2789.223877
Spike_AFFX-r2-Bs-lys_3-5-ratio3.793131
Spike_AFFX-r2-Bs-dap_M_signal105.523048
Spike_AFFX-r2-Bs-thr_M_detectionP
Probe Pair Threshold1
Spike_AFFX-r2-Bs-thr_3-5-ratio9.211758
Spike_AFFX-r2-Ec-bioB_avg-signal6.566241
Spike_AFFX-r2-Bs-thr_avg-signal35.244610
Spike_AFFX-r2-Bs-thr_5_detectionP
Spike_AFFX-r2-Bs-dap_3_detectionP
Spike_AFFX-r2-Ec-bioD_5_signal514.420471
Spike_AFFX-r2-Bs-phe_5_signal11.166242
Spike_AFFX-r2-P1-cre_5_detectionP
Spike_AFFX-r2-Ec-bioD_3_signal728.246643
RawQ3.276723
Spike_AFFX-r2-Bs-lys_5_signal11.844972
Signal(A)3.858927
%A37.268742
Signal(All)135.316986
Corner+Avg:219,Count:32
Spike_AFFX-r2-Ec-bioD_5_detectionP
Spike_AFFX-r2-Ec-bioC_3_signal142.065125
Spike_AFFX-r2-Bs-lys_3_detectionP
Spike_AFFX-r2-Bs-phe_3_signal26.743509
Spike_AFFX-r2-Ec-bioD_avg-signal621.333557
Spike_AFFX-r2-Bs-dap_5_detectionP
Spike_AFFX-r2-Bs-phe_5_detectionP
%P61.078476
Spike_AFFX-r2-Bs-lys_avg-signal22.514040
Spike_AFFX-r2-P1-cre_avg-signal2514.448730
Spike_AFFX-r2-Ec-bioB_M_detectionM
Spike_AFFX-r2-Bs-lys_M_detectionP
#Probe Sets Exceeding Probe Pair Threshold22810
%M1.652784
Spike_AFFX-r2-Bs-thr_3_signal71.458847
Spike_AFFX-r2-Ec-bioC_5_detectionP
Spike_AFFX-r2-Bs-phe_3_detectionM
Spike_AFFX-r2-Ec-bioC_3_detectionP
Spike_AFFX-r2-Bs-phe_avg-signal17.675554
Spike_AFFX-r2-Bs-dap_3_signal178.456070
Spike_AFFX-r2-Ec-bioD_3-5-ratio1.415664
Spike_AFFX-r2-Ec-bioD_3_detectionP
Spike_AFFX-r2-Ec-bioC_5_signal131.270737
#M377
Spike_AFFX-r2-Bs-lys_5_detectionP
Spike_AFFX-r2-Bs-lys_3_signal44.929523
Spike_AFFX-r2-Bs-dap_M_detectionP
Spike_AFFX-r2-Ec-bioC_3-5-ratio1.082230
Spike_AFFX-r2-Bs-thr_M_signal26.517626
Signal(P)218.783951
Spike_AFFX-r2-Bs-phe_3-5-ratio2.395032
Spike_AFFX-r2-Bs-thr_3_detectionP
Central-Avg:9413,Count:9
Spike_AFFX-r2-P1-cre_5_signal2239.673828
Spike_AFFX-r2-Bs-phe_M_detectionP
#A8501
Signal(M)15.060358
BackgroundAvg:84.00,Std:1.04,Min:81.5,Max:87.3
Spike_AFFX-r2-Ec-bioC_avg-signal136.667938
Spike_AFFX-r2-Bs-dap_avg-signal100.639137
Spike_AFFX-r2-Bs-dap_3-5-ratio9.948322
Spike_AFFX-r2-Ec-bioB_5_signal4.157687
ScaleMaskAll
BG2
VZ4
TGT100
Alpha10.040000
Alpha20.060000
SF0.359765
NF1.000000
HZ4
Tau0.015000


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