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Experiment: Cell type specific analysis of root epidermis

Experiment Description

NASCArrays Experiment Reference Number: NASCARRAYS-502

The experiment aims to analyse auxin signalling in root epidermis. We used two GFP marker lines (pCOBL9:GFP, pGL2:GFP) and fluorescent activated cell sorting to generate samples of trichoblast and atrichobast cells. Before protoplasting the roots were either mock treated or exposed to IAA (10µM) for 2 hours. Eventually the study will establish the topology of the auxin response system in these cell types and together with data from other sources will enable us to develop a model of auxin signalling in root epidermis. These samples represent a scaled down experiment aimed at testing the efficiency of our experimental protocol from cell sorting to microarray analysis.

About the Experimenter

Name:DR martin Kieffer
Head of Lab Name:DR Stefan Kepinski
Lab:
Address:Centre for plant sciences
University of Leeds
LEEDS
Postcode: LS29JT
Country: UK
 

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_treatment_design;
Number of Slides:7
 
Experimental Parameters:
Quality Control Measures Taken:
References:
 
Other Information:
titleCell type specific analysis of root epidermis

Slides in this Experiment

Hybridisation Set: Kieffer: Cell type specific analysis of root epidermis_genome

Slide: Kieffer_1-2_Atricoblast-mock_Rep2_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 ("Kieffer_1-2_Atricoblast-mock_Rep2_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: Kieffer_1-2_Atricoblast-mock_Rep2_ATH1
Organism: Arabidopsis thaliana
Alias: pGL2:GFP
Stock Code:
Genetic Background: Col-0
Age: 8 Days grown
Growth Conditions:
Stratification2 days of Cold treatment at 4C
Sterilisation70% ethanol 0.1%tween for 5 minutes 95% ethanol for 5 minutes
ProtocolSeeds grown in square petri dishes containing 70ml of MS medium at 22C under long-day condition
Percentage Agrose1
Substrate Sterilising Procedureautoclaving
Plant Spacinghigh density
Temperature22 °C average, 22 °C day, 22 °C night
Humiditytissue culture close to 90% % average, tissue culture close to 90% % day, tissue culture close to 90% % night
MediumMurashige & Skoog basal salt mixture. Modifications: + MS vitamins + 0.5g/l MES
Lighting
Developmental Stage:
Growth Stage(Source: Paradigm Genetics)1.03
Genetic Variation: T-DNA knock out mutation in GL2 promoter,At1g79840
Tissue: root tips
Additional Organism Information:
Sample DescriptionRoots from tissue cultured plants
Other Information:

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:5.335845
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-ratio1.124614
Spike_AFFX-r2-Ec-bioB_3-5-ratio1.147206
Spike_AFFX-r2-Bs-dap_5_signal183.061417
NoiseAvg:1.98,Std:0.08,Min:1.8,Max:2.2
Control DirectionAntisense
Spike_AFFX-r2-Ec-bioB_5_detectionP
Spike_AFFX-r2-Ec-bioB_3_detectionP
Spike_AFFX-r2-Bs-thr_5_signal163.652283
#P8055
Spike_AFFX-r2-Bs-phe_M_signal637.673401
Corner-Avg:11847,Count:32
Spike_AFFX-r2-P1-cre_3_detectionP
Spike_AFFX-r2-Ec-bioB_M_signal1157.528809
Spike_AFFX-r2-Ec-bioB_3_signal1377.745361
Spike_AFFX-r2-Bs-lys_M_signal1178.897339
Spike_AFFX-r2-P1-cre_3_signal51068.964844
Spike_AFFX-r2-Bs-lys_3-5-ratio11.998770
Spike_AFFX-r2-Bs-dap_M_signal4349.952637
Spike_AFFX-r2-Bs-thr_M_detectionP
Probe Pair Threshold1
Spike_AFFX-r2-Bs-thr_3-5-ratio74.359245
Spike_AFFX-r2-Ec-bioB_avg-signal1245.410278
Spike_AFFX-r2-Bs-thr_avg-signal4673.796875
Spike_AFFX-r2-Bs-thr_5_detectionP
Spike_AFFX-r2-Bs-dap_3_detectionP
Spike_AFFX-r2-Ec-bioD_5_signal12868.979492
Spike_AFFX-r2-Bs-phe_5_signal93.366203
Spike_AFFX-r2-P1-cre_5_detectionP
Spike_AFFX-r2-Ec-bioD_3_signal17575.451172
RawQ2.233613
Spike_AFFX-r2-Bs-lys_5_signal183.582504
Signal(A)23.493521
%A61.341518
Signal(All)168.150314
Corner+Avg:98,Count:32
Spike_AFFX-r2-Ec-bioD_5_detectionP
Spike_AFFX-r2-Ec-bioC_3_signal4118.296875
Spike_AFFX-r2-Bs-lys_3_detectionP
Spike_AFFX-r2-Bs-phe_3_signal3730.013184
Spike_AFFX-r2-Ec-bioD_avg-signal15222.214844
Spike_AFFX-r2-Bs-dap_5_detectionP
Spike_AFFX-r2-Bs-phe_5_detectionP
%P35.313457
Spike_AFFX-r2-Bs-lys_avg-signal1188.414673
Spike_AFFX-r2-P1-cre_avg-signal48239.593750
Spike_AFFX-r2-Ec-bioB_M_detectionP
Spike_AFFX-r2-Bs-lys_M_detectionP
#Probe Sets Exceeding Probe Pair Threshold22810
%M3.345024
Spike_AFFX-r2-Bs-thr_3_signal12169.060547
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-signal1487.017578
Spike_AFFX-r2-Bs-dap_3_signal15405.951172
Spike_AFFX-r2-Ec-bioD_3-5-ratio1.365722
Spike_AFFX-r2-Ec-bioD_3_detectionP
Spike_AFFX-r2-Ec-bioC_5_signal3564.492676
#M763
Spike_AFFX-r2-Bs-lys_5_detectionP
Spike_AFFX-r2-Bs-lys_3_signal2202.764160
Spike_AFFX-r2-Bs-dap_M_detectionP
Spike_AFFX-r2-Ec-bioC_3-5-ratio1.155367
Spike_AFFX-r2-Bs-thr_M_signal1688.677856
Signal(P)427.963501
Spike_AFFX-r2-Bs-phe_3-5-ratio39.950356
Spike_AFFX-r2-Bs-thr_3_detectionP
Central-Avg:9434,Count:9
Spike_AFFX-r2-P1-cre_5_signal45410.222656
Spike_AFFX-r2-Bs-phe_M_detectionP
#A13992
Signal(M)78.035866
BackgroundAvg:51.98,Std:0.71,Min:50.4,Max:54.4
Spike_AFFX-r2-Ec-bioC_avg-signal3841.394775
Spike_AFFX-r2-Bs-dap_avg-signal6646.321777
Spike_AFFX-r2-Bs-dap_3-5-ratio84.157280
Spike_AFFX-r2-Ec-bioB_5_signal1200.956909
ScaleMaskAll
BG2
VZ4
TGT100
Alpha10.040000
Alpha20.060000
SF5.335845
NF1.000000
HZ4
Tau0.015000

Slide: Kieffer_1-1_Atricoblast-mock_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 ("Kieffer_1-1_Atricoblast-mock_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: Kieffer_1-1_Atricoblast-mock_Rep1_ATH1
Organism: Arabidopsis thaliana
Alias: pGL2:GFP
Stock Code:
Genetic Background: Col-0
Growth Conditions:
Stratification2 days of Cold treatment at 4C
Sterilisation70% ethanol 0.1%tween for 5 minutes 95% ethanol for 5 minutes
ProtocolSeeds grown in square petri dishes containing 70ml of MS medium at 22C under long-day condition
Percentage Agrose1
Substrate Sterilising Procedureautoclaving
Plant Spacinghigh density
Temperature22 °C average, 22 °C day, 22 °C night
Humiditytissue culture close to 90% % average, tissue culture close to 90% % day, tissue culture close to 90% % night
MediumMurashige & Skoog basal salt mixture. Modifications: + MS vitamins + 0.5g/l MES
Lighting
Developmental Stage:
Growth Stage(Source: Paradigm Genetics)1.03
Genetic Variation: T-DNA knock out mutation in GL2 promoter,At1g79840
Tissue: root tips
Additional Organism Information:
Sample DescriptionRoots from tissue cultured plants
Other Information:

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:6.642985
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-ratio1.167466
Spike_AFFX-r2-Ec-bioB_3-5-ratio1.179543
Spike_AFFX-r2-Bs-dap_5_signal482.018250
NoiseAvg:2.02,Std:0.07,Min:1.9,Max:2.3
Control DirectionAntisense
Spike_AFFX-r2-Ec-bioB_5_detectionP
Spike_AFFX-r2-Ec-bioB_3_detectionP
Spike_AFFX-r2-Bs-thr_5_signal237.119995
#P6930
Spike_AFFX-r2-Bs-phe_M_signal1405.124634
Corner-Avg:12138,Count:32
Spike_AFFX-r2-P1-cre_3_detectionP
Spike_AFFX-r2-Ec-bioB_M_signal1882.083252
Spike_AFFX-r2-Ec-bioB_3_signal1686.100586
Spike_AFFX-r2-Bs-lys_M_signal1722.347290
Spike_AFFX-r2-P1-cre_3_signal65825.031250
Spike_AFFX-r2-Bs-lys_3-5-ratio23.792622
Spike_AFFX-r2-Bs-dap_M_signal7956.721191
Spike_AFFX-r2-Bs-thr_M_detectionP
Probe Pair Threshold1
Spike_AFFX-r2-Bs-thr_3-5-ratio101.008469
Spike_AFFX-r2-Ec-bioB_avg-signal1665.878540
Spike_AFFX-r2-Bs-thr_avg-signal8962.743164
Spike_AFFX-r2-Bs-thr_5_detectionP
Spike_AFFX-r2-Bs-dap_3_detectionP
Spike_AFFX-r2-Ec-bioD_5_signal16931.593750
Spike_AFFX-r2-Bs-phe_5_signal244.438675
Spike_AFFX-r2-P1-cre_5_detectionP
Spike_AFFX-r2-Ec-bioD_3_signal19395.894531
RawQ2.213341
Spike_AFFX-r2-Bs-lys_5_signal172.281937
Signal(A)25.945841
%A66.865410
Signal(All)175.822571
Corner+Avg:103,Count:32
Spike_AFFX-r2-Ec-bioD_5_detectionP
Spike_AFFX-r2-Ec-bioC_3_signal6219.229004
Spike_AFFX-r2-Bs-lys_3_detectionP
Spike_AFFX-r2-Bs-phe_3_signal9513.234375
Spike_AFFX-r2-Ec-bioD_avg-signal18163.744141
Spike_AFFX-r2-Bs-dap_5_detectionP
Spike_AFFX-r2-Bs-phe_5_detectionP
%P30.381413
Spike_AFFX-r2-Bs-lys_avg-signal1997.889282
Spike_AFFX-r2-P1-cre_avg-signal61103.937500
Spike_AFFX-r2-Ec-bioB_M_detectionP
Spike_AFFX-r2-Bs-lys_M_detectionP
#Probe Sets Exceeding Probe Pair Threshold22810
%M2.753178
Spike_AFFX-r2-Bs-thr_3_signal23951.126953
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-signal3720.932617
Spike_AFFX-r2-Bs-dap_3_signal20609.683594
Spike_AFFX-r2-Ec-bioD_3-5-ratio1.145545
Spike_AFFX-r2-Ec-bioD_3_detectionP
Spike_AFFX-r2-Ec-bioC_5_signal4940.760742
#M628
Spike_AFFX-r2-Bs-lys_5_detectionP
Spike_AFFX-r2-Bs-lys_3_signal4099.039063
Spike_AFFX-r2-Bs-dap_M_detectionP
Spike_AFFX-r2-Ec-bioC_3-5-ratio1.258759
Spike_AFFX-r2-Bs-thr_M_signal2699.983887
Signal(P)513.431030
Spike_AFFX-r2-Bs-phe_3-5-ratio38.918697
Spike_AFFX-r2-Bs-thr_3_detectionP
Central-Avg:9354,Count:9
Spike_AFFX-r2-P1-cre_5_signal56382.839844
Spike_AFFX-r2-Bs-phe_M_detectionP
#A15252
Signal(M)90.302299
BackgroundAvg:51.86,Std:1.06,Min:50.0,Max:55.7
Spike_AFFX-r2-Ec-bioC_avg-signal5579.995117
Spike_AFFX-r2-Bs-dap_avg-signal9682.807617
Spike_AFFX-r2-Bs-dap_3-5-ratio42.757061
Spike_AFFX-r2-Ec-bioB_5_signal1429.452148
ScaleMaskAll
BG2
VZ4
TGT100
Alpha10.040000
Alpha20.060000
SF6.642985
NF1.000000
HZ4
Tau0.015000

Slide: Kieffer_1-3_Atricoblast-auxin_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 ("Kieffer_1-3_Atricoblast-auxin_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: Kieffer_1-3_Atricoblast-auxin_Rep1_ATH1
Organism: Arabidopsis thaliana
Alias: pGL2:GFP
Stock Code:
Genetic Background: Col-0
Age: 8 Days grown
Growth Conditions:
Stratification2 days of Cold treatment at 4C
Sterilisation70% ethanol 0.1%tween for 5 minutes 95% ethanol for 5 minutes
ProtocolSeeds grown in square petri dishes containing 70ml of MS medium at 22C under long-day condition
Percentage Agrose1
Substrate Sterilising Procedureautoclaving
Plant Spacinghigh density
Temperature22 °C average, 22 °C day, 22 °C night
Humiditytissue culture close to 90% % average, tissue culture close to 90% % day, tissue culture close to 90% % night
MediumMurashige & Skoog basal salt mixture. Modifications: + MS vitamins + 0.5g/l MES
Lighting
Developmental Stage:
Growth Stage(Source: Paradigm Genetics)1.03
Genetic Variation: T-DNA knock out mutation in GL2 promoter,At1g79840
Tissue: root tips
in vivo Treatment: IAA 1OµM for 2hrs
Additional Organism Information:
Sample DescriptionRoots from tissue cultured plants
Other Information:

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:5.655102
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-ratio1.095257
Spike_AFFX-r2-Ec-bioB_3-5-ratio0.626809
Spike_AFFX-r2-Bs-dap_5_signal36.301098
NoiseAvg:1.73,Std:0.05,Min:1.6,Max:1.9
Control DirectionAntisense
Spike_AFFX-r2-Ec-bioB_5_detectionP
Spike_AFFX-r2-Ec-bioB_3_detectionP
Spike_AFFX-r2-Bs-thr_5_signal32.643192
#P7353
Spike_AFFX-r2-Bs-phe_M_signal160.687698
Corner-Avg:10149,Count:32
Spike_AFFX-r2-P1-cre_3_detectionP
Spike_AFFX-r2-Ec-bioB_M_signal975.692932
Spike_AFFX-r2-Ec-bioB_3_signal619.588196
Spike_AFFX-r2-Bs-lys_M_signal110.081451
Spike_AFFX-r2-P1-cre_3_signal37506.277344
Spike_AFFX-r2-Bs-lys_3-5-ratio360.983856
Spike_AFFX-r2-Bs-dap_M_signal777.477783
Spike_AFFX-r2-Bs-thr_M_detectionP
Probe Pair Threshold1
Spike_AFFX-r2-Bs-thr_3-5-ratio102.477295
Spike_AFFX-r2-Ec-bioB_avg-signal861.253723
Spike_AFFX-r2-Bs-thr_avg-signal1248.166870
Spike_AFFX-r2-Bs-thr_5_detectionA
Spike_AFFX-r2-Bs-dap_3_detectionP
Spike_AFFX-r2-Ec-bioD_5_signal8543.791016
Spike_AFFX-r2-Bs-phe_5_signal4.782137
Spike_AFFX-r2-P1-cre_5_detectionP
Spike_AFFX-r2-Ec-bioD_3_signal11717.712891
RawQ1.967621
Spike_AFFX-r2-Bs-lys_5_signal1.093431
Signal(A)22.466667
%A65.124947
Signal(All)168.152222
Corner+Avg:81,Count:32
Spike_AFFX-r2-Ec-bioD_5_detectionP
Spike_AFFX-r2-Ec-bioC_3_signal3948.934814
Spike_AFFX-r2-Bs-lys_3_detectionP
Spike_AFFX-r2-Bs-phe_3_signal1064.912964
Spike_AFFX-r2-Ec-bioD_avg-signal10130.751953
Spike_AFFX-r2-Bs-dap_5_detectionA
Spike_AFFX-r2-Bs-phe_5_detectionA
%P32.235863
Spike_AFFX-r2-Bs-lys_avg-signal168.628571
Spike_AFFX-r2-P1-cre_avg-signal35875.273438
Spike_AFFX-r2-Ec-bioB_M_detectionP
Spike_AFFX-r2-Bs-lys_M_detectionP
#Probe Sets Exceeding Probe Pair Threshold22810
%M2.639193
Spike_AFFX-r2-Bs-thr_3_signal3345.186035
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-signal410.127594
Spike_AFFX-r2-Bs-dap_3_signal5231.992676
Spike_AFFX-r2-Ec-bioD_3-5-ratio1.371489
Spike_AFFX-r2-Ec-bioD_3_detectionP
Spike_AFFX-r2-Ec-bioC_5_signal3354.705811
#M602
Spike_AFFX-r2-Bs-lys_5_detectionA
Spike_AFFX-r2-Bs-lys_3_signal394.710815
Spike_AFFX-r2-Bs-dap_M_detectionP
Spike_AFFX-r2-Ec-bioC_3-5-ratio1.177133
Spike_AFFX-r2-Bs-thr_M_signal366.671478
Signal(P)469.790833
Spike_AFFX-r2-Bs-phe_3-5-ratio222.685593
Spike_AFFX-r2-Bs-thr_3_detectionP
Central-Avg:8064,Count:9
Spike_AFFX-r2-P1-cre_5_signal34244.273438
Spike_AFFX-r2-Bs-phe_M_detectionP
#A14855
Signal(M)78.800262
BackgroundAvg:69.87,Std:0.74,Min:68.2,Max:72.3
Spike_AFFX-r2-Ec-bioC_avg-signal3651.820313
Spike_AFFX-r2-Bs-dap_avg-signal2015.257324
Spike_AFFX-r2-Bs-dap_3-5-ratio144.127670
Spike_AFFX-r2-Ec-bioB_5_signal988.479980
ScaleMaskAll
BG2
VZ4
TGT100
Alpha10.040000
Alpha20.060000
SF5.655102
NF1.000000
HZ4
Tau0.015000

Slide: Kieffer_1-4_Atricoblast-auxin_Rep2_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 ("Kieffer_1-4_Atricoblast-auxin_Rep2_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: Kieffer_1-4_Atricoblast-auxin_Rep2_ATH1
Organism: Arabidopsis thaliana
Alias: pGL2:GFP
Stock Code:
Genetic Background: Col-0
Age: 8 Days grown
Growth Conditions:
Stratification2 days of Cold treatment at 4C
Sterilisation70% ethanol 0.1%tween for 5 minutes 95% ethanol for 5 minutes
ProtocolSeeds grown in square petri dishes containing 70ml of MS medium at 22C under long-day condition
Percentage Agrose1
Substrate Sterilising Procedureautoclaving
Plant Spacinghigh density
Temperature22 °C average, 22 °C day, 22 °C night
Humiditytissue culture close to 90% % average, tissue culture close to 90% % day, tissue culture close to 90% % night
MediumMurashige & Skoog basal salt mixture. Modifications: + MS vitamins + 0.5g/l MES
Lighting
Developmental Stage:
Growth Stage(Source: Paradigm Genetics)1.03
Genetic Variation: T-DNA knock out mutation in GL2 promoter,At1g79840
Tissue: root tips
in vivo Treatment: IAA 1OµM for 2hrs
Additional Organism Information:
Sample DescriptionRoots from tissue cultured plants
Other Information:

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:17.133579
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-ratio1.150454
Spike_AFFX-r2-Ec-bioB_3-5-ratio1.287755
Spike_AFFX-r2-Bs-dap_5_signal1378.387817
NoiseAvg:2.01,Std:0.27,Min:1.7,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_signal1305.893677
#P3377
Spike_AFFX-r2-Bs-phe_M_signal4431.807617
Corner-Avg:12478,Count:32
Spike_AFFX-r2-P1-cre_3_detectionP
Spike_AFFX-r2-Ec-bioB_M_signal5874.498535
Spike_AFFX-r2-Ec-bioB_3_signal5746.523438
Spike_AFFX-r2-Bs-lys_M_signal6050.042480
Spike_AFFX-r2-P1-cre_3_signal191811.359375
Spike_AFFX-r2-Bs-lys_3-5-ratio15.862544
Spike_AFFX-r2-Bs-dap_M_signal19771.675781
Spike_AFFX-r2-Bs-thr_M_detectionP
Probe Pair Threshold1
Spike_AFFX-r2-Bs-thr_3-5-ratio46.331600
Spike_AFFX-r2-Ec-bioB_avg-signal5361.152832
Spike_AFFX-r2-Bs-thr_avg-signal24037.587891
Spike_AFFX-r2-Bs-thr_5_detectionP
Spike_AFFX-r2-Bs-dap_3_detectionP
Spike_AFFX-r2-Ec-bioD_5_signal63495.656250
Spike_AFFX-r2-Bs-phe_5_signal733.259766
Spike_AFFX-r2-P1-cre_5_detectionP
Spike_AFFX-r2-Ec-bioD_3_signal73739.476563
RawQ2.185771
Spike_AFFX-r2-Bs-lys_5_signal648.330627
Signal(A)55.297401
%A82.801399
Signal(All)208.412262
Corner+Avg:104,Count:32
Spike_AFFX-r2-Ec-bioD_5_detectionP
Spike_AFFX-r2-Ec-bioC_3_signal19295.271484
Spike_AFFX-r2-Bs-lys_3_detectionP
Spike_AFFX-r2-Bs-phe_3_signal24309.339844
Spike_AFFX-r2-Ec-bioD_avg-signal68617.562500
Spike_AFFX-r2-Bs-dap_5_detectionP
Spike_AFFX-r2-Bs-phe_5_detectionP
%P14.804910
Spike_AFFX-r2-Bs-lys_avg-signal5660.848145
Spike_AFFX-r2-P1-cre_avg-signal179268.984375
Spike_AFFX-r2-Ec-bioB_M_detectionP
Spike_AFFX-r2-Bs-lys_M_detectionP
#Probe Sets Exceeding Probe Pair Threshold22810
%M2.393687
Spike_AFFX-r2-Bs-thr_3_signal60504.144531
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-signal9824.802734
Spike_AFFX-r2-Bs-dap_3_signal63779.308594
Spike_AFFX-r2-Ec-bioD_3-5-ratio1.161331
Spike_AFFX-r2-Ec-bioD_3_detectionP
Spike_AFFX-r2-Ec-bioC_5_signal15833.220703
#M546
Spike_AFFX-r2-Bs-lys_5_detectionP
Spike_AFFX-r2-Bs-lys_3_signal10284.172852
Spike_AFFX-r2-Bs-dap_M_detectionP
Spike_AFFX-r2-Ec-bioC_3-5-ratio1.218657
Spike_AFFX-r2-Bs-thr_M_signal10302.728516
Signal(P)1066.542480
Spike_AFFX-r2-Bs-phe_3-5-ratio33.152424
Spike_AFFX-r2-Bs-thr_3_detectionP
Central-Avg:10248,Count:9
Spike_AFFX-r2-P1-cre_5_signal166726.609375
Spike_AFFX-r2-Bs-phe_M_detectionP
#A18887
Signal(M)197.376648
BackgroundAvg:51.36,Std:0.75,Min:49.8,Max:53.3
Spike_AFFX-r2-Ec-bioC_avg-signal17564.246094
Spike_AFFX-r2-Bs-dap_avg-signal28309.791016
Spike_AFFX-r2-Bs-dap_3-5-ratio46.270947
Spike_AFFX-r2-Ec-bioB_5_signal4462.436523
ScaleMaskAll
BG2
VZ4
TGT100
Alpha10.040000
Alpha20.060000
SF17.133579
NF1.000000
HZ4
Tau0.015000

Slide: Kieffer_1-5_Trichoblast-mock_Rep1_ATH1

Add to Slide Selection

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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 ("Kieffer_1-5_Trichoblast-mock_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: Kieffer_1-5_Trichoblast-mock_Rep1_ATH1
Organism: Arabidopsis thaliana
Alias: pCOBL9:GFP
Stock Code:
Genetic Background: Col-0
Age: 8 Days grown
Growth Conditions:
Stratification2 days of Cold treatment at 4C
Sterilisation70% ethanol 0.1%tween for 5 minutes 95% ethanol for 5 minutes
ProtocolSeeds grown in square petri dishes containing 70ml of MS medium at 22C under long-day condition
Percentage Agrose1
Substrate Sterilising Procedureautoclaving
Plant Spacinghigh density
Temperature22 °C average, 22 °C day, 22 °C night
Humiditytissue culture close to 90% % average, tissue culture close to 90% % day, tissue culture close to 90% % night
MediumMurashige & Skoog basal salt mixture. Modifications: + MS vitamins + 0.5g/l MES
Lighting
Developmental Stage:
Growth Stage(Source: Paradigm Genetics)1.03
Genetic Variation: T-DNA knock out mutation in COBL9 promoter,At5g49270
Tissue: root tips
Additional Organism Information:
Sample DescriptionRoots from tissue cultured plants
Other Information:

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:8.805079
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-ratio1.174058
Spike_AFFX-r2-Ec-bioB_3-5-ratio1.064739
Spike_AFFX-r2-Bs-dap_5_signal375.296692
NoiseAvg:2.02,Std:0.09,Min:1.8,Max:2.3
Control DirectionAntisense
Spike_AFFX-r2-Ec-bioB_5_detectionP
Spike_AFFX-r2-Ec-bioB_3_detectionP
Spike_AFFX-r2-Bs-thr_5_signal337.616974
#P6122
Spike_AFFX-r2-Bs-phe_M_signal1116.975830
Corner-Avg:11745,Count:32
Spike_AFFX-r2-P1-cre_3_detectionP
Spike_AFFX-r2-Ec-bioB_M_signal2080.604248
Spike_AFFX-r2-Ec-bioB_3_signal1901.473022
Spike_AFFX-r2-Bs-lys_M_signal1237.858643
Spike_AFFX-r2-P1-cre_3_signal77205.945313
Spike_AFFX-r2-Bs-lys_3-5-ratio25.164803
Spike_AFFX-r2-Bs-dap_M_signal6044.862305
Spike_AFFX-r2-Bs-thr_M_detectionP
Probe Pair Threshold1
Spike_AFFX-r2-Bs-thr_3-5-ratio61.099224
Spike_AFFX-r2-Ec-bioB_avg-signal1922.644897
Spike_AFFX-r2-Bs-thr_avg-signal7775.912109
Spike_AFFX-r2-Bs-thr_5_detectionP
Spike_AFFX-r2-Bs-dap_3_detectionP
Spike_AFFX-r2-Ec-bioD_5_signal19289.687500
Spike_AFFX-r2-Bs-phe_5_signal174.708466
Spike_AFFX-r2-P1-cre_5_detectionP
Spike_AFFX-r2-Ec-bioD_3_signal25255.355469
RawQ2.297333
Spike_AFFX-r2-Bs-lys_5_signal98.119537
Signal(A)34.447624
%A70.241119
Signal(All)174.622513
Corner+Avg:91,Count:32
Spike_AFFX-r2-Ec-bioD_5_detectionP
Spike_AFFX-r2-Ec-bioC_3_signal7072.552246
Spike_AFFX-r2-Bs-lys_3_detectionP
Spike_AFFX-r2-Bs-phe_3_signal6092.153320
Spike_AFFX-r2-Ec-bioD_avg-signal22272.521484
Spike_AFFX-r2-Bs-dap_5_detectionP
Spike_AFFX-r2-Bs-phe_5_detectionP
%P26.839106
Spike_AFFX-r2-Bs-lys_avg-signal1268.379028
Spike_AFFX-r2-P1-cre_avg-signal71482.929688
Spike_AFFX-r2-Ec-bioB_M_detectionP
Spike_AFFX-r2-Bs-lys_M_detectionP
#Probe Sets Exceeding Probe Pair Threshold22810
%M2.919772
Spike_AFFX-r2-Bs-thr_3_signal20628.134766
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-signal2461.279053
Spike_AFFX-r2-Bs-dap_3_signal22186.755859
Spike_AFFX-r2-Ec-bioD_3-5-ratio1.309267
Spike_AFFX-r2-Ec-bioD_3_detectionP
Spike_AFFX-r2-Ec-bioC_5_signal5813.978516
#M666
Spike_AFFX-r2-Bs-lys_5_detectionP
Spike_AFFX-r2-Bs-lys_3_signal2469.158691
Spike_AFFX-r2-Bs-dap_M_detectionP
Spike_AFFX-r2-Ec-bioC_3-5-ratio1.216474
Spike_AFFX-r2-Bs-thr_M_signal2361.984375
Signal(P)547.833191
Spike_AFFX-r2-Bs-phe_3-5-ratio34.870396
Spike_AFFX-r2-Bs-thr_3_detectionP
Central-Avg:9120,Count:9
Spike_AFFX-r2-P1-cre_5_signal65759.914063
Spike_AFFX-r2-Bs-phe_M_detectionP
#A16022
Signal(M)116.193802
BackgroundAvg:55.17,Std:1.36,Min:52.8,Max:59.7
Spike_AFFX-r2-Ec-bioC_avg-signal6443.265625
Spike_AFFX-r2-Bs-dap_avg-signal9535.637695
Spike_AFFX-r2-Bs-dap_3-5-ratio59.117908
Spike_AFFX-r2-Ec-bioB_5_signal1785.857666
ScaleMaskAll
BG2
VZ4
TGT100
Alpha10.040000
Alpha20.060000
SF8.805079
NF1.000000
HZ4
Tau0.015000

Slide: Kieffer_1-6_Trichoblast-mock_Rep2_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 ("Kieffer_1-6_Trichoblast-mock_Rep2_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: Kieffer_1-6_Trichoblast-mock_Rep2_ATH1
Organism: Arabidopsis thaliana
Alias: pCOBL9:GFP
Stock Code:
Genetic Background: Col-0
Age: 8 Days grown
Growth Conditions:
Stratification2 days of Cold treatment at 4C
Sterilisation70% ethanol 0.1%tween for 5 minutes 95% ethanol for 5 minutes
ProtocolSeeds grown in square petri dishes containing 70ml of MS medium at 22C under long-day condition
Percentage Agrose1
Substrate Sterilising Procedureautoclaving
Plant Spacinghigh density
Temperature22 °C average, 22 °C day, 22 °C night
Humiditytissue culture close to 90% % average, tissue culture close to 90% % day, tissue culture close to 90% % night
MediumMurashige & Skoog basal salt mixture. Modifications: + MS vitamins + 0.5g/l MES
Lighting
Developmental Stage:
Growth Stage(Source: Paradigm Genetics)1.03
Genetic Variation: T-DNA knock out mutation in COBL9 promoter,At5g49270
Tissue: root tips
Additional Organism Information:
Sample DescriptionRoots from tissue cultured plants
Other Information:

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:3.44508
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-ratio1.193311
Spike_AFFX-r2-Ec-bioB_3-5-ratio0.927936
Spike_AFFX-r2-Bs-dap_5_signal56.313751
NoiseAvg:1.92,Std:0.07,Min:1.7,Max:2.1
Control DirectionAntisense
Spike_AFFX-r2-Ec-bioB_5_detectionP
Spike_AFFX-r2-Ec-bioB_3_detectionP
Spike_AFFX-r2-Bs-thr_5_signal64.140419
#P9837
Spike_AFFX-r2-Bs-phe_M_signal243.767792
Corner-Avg:12759,Count:32
Spike_AFFX-r2-P1-cre_3_detectionP
Spike_AFFX-r2-Ec-bioB_M_signal723.922974
Spike_AFFX-r2-Ec-bioB_3_signal677.882935
Spike_AFFX-r2-Bs-lys_M_signal437.742645
Spike_AFFX-r2-P1-cre_3_signal31368.375000
Spike_AFFX-r2-Bs-lys_3-5-ratio13.166974
Spike_AFFX-r2-Bs-dap_M_signal2251.376953
Spike_AFFX-r2-Bs-thr_M_detectionP
Probe Pair Threshold1
Spike_AFFX-r2-Bs-thr_3-5-ratio88.787743
Spike_AFFX-r2-Ec-bioB_avg-signal710.777893
Spike_AFFX-r2-Bs-thr_avg-signal2163.208252
Spike_AFFX-r2-Bs-thr_5_detectionP
Spike_AFFX-r2-Bs-dap_3_detectionP
Spike_AFFX-r2-Ec-bioD_5_signal6645.833984
Spike_AFFX-r2-Bs-phe_5_signal37.536182
Spike_AFFX-r2-P1-cre_5_detectionP
Spike_AFFX-r2-Ec-bioD_3_signal9435.574219
RawQ2.168345
Spike_AFFX-r2-Bs-lys_5_signal71.130402
Signal(A)15.226842
%A53.958790
Signal(All)159.421570
Corner+Avg:101,Count:32
Spike_AFFX-r2-Ec-bioD_5_detectionP
Spike_AFFX-r2-Ec-bioC_3_signal2407.786865
Spike_AFFX-r2-Bs-lys_3_detectionP
Spike_AFFX-r2-Bs-phe_3_signal2305.937744
Spike_AFFX-r2-Ec-bioD_avg-signal8040.704102
Spike_AFFX-r2-Bs-dap_5_detectionP
Spike_AFFX-r2-Bs-phe_5_detectionP
%P43.125820
Spike_AFFX-r2-Bs-lys_avg-signal481.815063
Spike_AFFX-r2-P1-cre_avg-signal28827.611328
Spike_AFFX-r2-Ec-bioB_M_detectionP
Spike_AFFX-r2-Bs-lys_M_detectionP
#Probe Sets Exceeding Probe Pair Threshold22810
%M2.915388
Spike_AFFX-r2-Bs-thr_3_signal5694.882813
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-signal862.413879
Spike_AFFX-r2-Bs-dap_3_signal9100.959961
Spike_AFFX-r2-Ec-bioD_3-5-ratio1.419773
Spike_AFFX-r2-Ec-bioD_3_detectionP
Spike_AFFX-r2-Ec-bioC_5_signal2029.371582
#M665
Spike_AFFX-r2-Bs-lys_5_detectionP
Spike_AFFX-r2-Bs-lys_3_signal936.572144
Spike_AFFX-r2-Bs-dap_M_detectionP
Spike_AFFX-r2-Ec-bioC_3-5-ratio1.186469
Spike_AFFX-r2-Bs-thr_M_signal730.601257
Signal(P)347.041656
Spike_AFFX-r2-Bs-phe_3-5-ratio61.432400
Spike_AFFX-r2-Bs-thr_3_detectionP
Central-Avg:10200,Count:9
Spike_AFFX-r2-P1-cre_5_signal26286.847656
Spike_AFFX-r2-Bs-phe_M_detectionP
#A12308
Signal(M)52.849998
BackgroundAvg:49.41,Std:1.16,Min:47.8,Max:53.1
Spike_AFFX-r2-Ec-bioC_avg-signal2218.579102
Spike_AFFX-r2-Bs-dap_avg-signal3802.883545
Spike_AFFX-r2-Bs-dap_3-5-ratio161.611679
Spike_AFFX-r2-Ec-bioB_5_signal730.527832
ScaleMaskAll
BG2
VZ4
TGT100
Alpha10.040000
Alpha20.060000
SF3.445080
NF1.000000
HZ4
Tau0.015000

Slide: Kieffer_1-7_Trichoblast-auxin_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 ("Kieffer_1-7_Trichoblast-auxin_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: Kieffer_1-7_Trichoblast-auxin_Rep1_ATH1
Organism: Arabidopsis thaliana
Alias: pCOBL9:GFP
Stock Code:
Genetic Background: Col-0
Age: 8 Days grown
Growth Conditions:
Stratification2 days of Cold treatment at 4C
Sterilisation70% ethanol 0.1%tween for 5 minutes 95% ethanol for 5 minutes
ProtocolSeeds grown in square petri dishes containing 70ml of MS medium at 22C under long-day condition
Percentage Agrose1
Substrate Sterilising Procedureautoclaving
Plant Spacinghigh density
Temperature22 °C average, 22 °C day, 22 °C night
Humiditytissue culture close to 90% % average, tissue culture close to 90% % day, tissue culture close to 90% % night
MediumMurashige & Skoog basal salt mixture. Modifications: + MS vitamins + 0.5g/l MES
Lighting
Developmental Stage:
Growth Stage(Source: Paradigm Genetics)1.03
Genetic Variation: T-DNA knock out mutation in COBL9 promoter,At5g49270
Tissue: root tips
in vivo Treatment: IAA 1OµM for 2hrs
Additional Organism Information:
Sample DescriptionRoots from tissue cultured plants
Other Information:

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:2.00824
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-ratio1.290633
Spike_AFFX-r2-Ec-bioB_3-5-ratio0.797495
Spike_AFFX-r2-Bs-dap_5_signal90.594704
NoiseAvg:2.30,Std:0.08,Min:2.1,Max:2.6
Control DirectionAntisense
Spike_AFFX-r2-Ec-bioB_5_detectionP
Spike_AFFX-r2-Ec-bioB_3_detectionP
Spike_AFFX-r2-Bs-thr_5_signal78.920578
#P10967
Spike_AFFX-r2-Bs-phe_M_signal209.996872
Corner-Avg:11720,Count:32
Spike_AFFX-r2-P1-cre_3_detectionP
Spike_AFFX-r2-Ec-bioB_M_signal294.584320
Spike_AFFX-r2-Ec-bioB_3_signal235.480972
Spike_AFFX-r2-Bs-lys_M_signal245.134872
Spike_AFFX-r2-P1-cre_3_signal16005.452148
Spike_AFFX-r2-Bs-lys_3-5-ratio12.712565
Spike_AFFX-r2-Bs-dap_M_signal1621.669312
Spike_AFFX-r2-Bs-thr_M_detectionP
Probe Pair Threshold1
Spike_AFFX-r2-Bs-thr_3-5-ratio54.207298
Spike_AFFX-r2-Ec-bioB_avg-signal275.113739
Spike_AFFX-r2-Bs-thr_avg-signal1651.239746
Spike_AFFX-r2-Bs-thr_5_detectionP
Spike_AFFX-r2-Bs-dap_3_detectionP
Spike_AFFX-r2-Ec-bioD_5_signal2535.966064
Spike_AFFX-r2-Bs-phe_5_signal40.425606
Spike_AFFX-r2-P1-cre_5_detectionP
Spike_AFFX-r2-Ec-bioD_3_signal3866.132080
RawQ2.330051
Spike_AFFX-r2-Bs-lys_5_signal44.110947
Signal(A)11.242834
%A49.188953
Signal(All)154.915741
Corner+Avg:99,Count:32
Spike_AFFX-r2-Ec-bioD_5_detectionP
Spike_AFFX-r2-Ec-bioC_3_signal955.415222
Spike_AFFX-r2-Bs-lys_3_detectionP
Spike_AFFX-r2-Bs-phe_3_signal1322.468140
Spike_AFFX-r2-Ec-bioD_avg-signal3201.049072
Spike_AFFX-r2-Bs-dap_5_detectionP
Spike_AFFX-r2-Bs-phe_5_detectionP
%P48.079788
Spike_AFFX-r2-Bs-lys_avg-signal283.336395
Spike_AFFX-r2-P1-cre_avg-signal14203.345703
Spike_AFFX-r2-Ec-bioB_M_detectionP
Spike_AFFX-r2-Bs-lys_M_detectionP
#Probe Sets Exceeding Probe Pair Threshold22810
%M2.731258
Spike_AFFX-r2-Bs-thr_3_signal4278.071289
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-signal524.296875
Spike_AFFX-r2-Bs-dap_3_signal4153.046875
Spike_AFFX-r2-Ec-bioD_3-5-ratio1.524520
Spike_AFFX-r2-Ec-bioD_3_detectionP
Spike_AFFX-r2-Ec-bioC_5_signal687.173523
#M623
Spike_AFFX-r2-Bs-lys_5_detectionP
Spike_AFFX-r2-Bs-lys_3_signal560.763306
Spike_AFFX-r2-Bs-dap_M_detectionP
Spike_AFFX-r2-Ec-bioC_3-5-ratio1.390355
Spike_AFFX-r2-Bs-thr_M_signal596.727661
Signal(P)308.611328
Spike_AFFX-r2-Bs-phe_3-5-ratio32.713627
Spike_AFFX-r2-Bs-thr_3_detectionP
Central-Avg:8225,Count:9
Spike_AFFX-r2-P1-cre_5_signal12401.239258
Spike_AFFX-r2-Bs-phe_M_detectionP
#A11220
Signal(M)36.826385
BackgroundAvg:58.61,Std:0.57,Min:57.3,Max:61.0
Spike_AFFX-r2-Ec-bioC_avg-signal821.294373
Spike_AFFX-r2-Bs-dap_avg-signal1955.103638
Spike_AFFX-r2-Bs-dap_3-5-ratio45.842049
Spike_AFFX-r2-Ec-bioB_5_signal295.275909
ScaleMaskAll
BG2
VZ4
TGT100
Alpha10.040000
Alpha20.060000
SF2.008240
NF1.000000
HZ4
Tau0.015000


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