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Experiment: Gene expression in root tips of curly leaf mutants

Experiment Description

NASCArrays Experiment Reference Number: NASCARRAYS-617

Our group would like to compare gene expression between wildtype and curly leaf (clf) mutant roots. Wildtype and mutant plants were grown vertically in MS agar and the root tips harvested after 5 days. RNA was extracted using RNEasy Plant mini kit.

About the Experimenter

Name:Dr Corina Belle Villar
Head of Lab Name:Dr Claudia Koehler
Lab:
Address:Institute of Plant Sciences
Plant Developmental Biology
ETH Zurich LFW E33.1
Universitaetstrasse 2
Postcode: 8092 Zurich
Country: Switzerland
 

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: genetic_modification_design
Number of Slides:6
 
Experimental Parameters:
parametergene_knock_out
Quality Control Measures Taken:
References:
 
Other Information:

Slides in this Experiment

Hybridisation Set: Villar: Gene expression in root tips of curly leaf mutants

Slide: Villar_617-1_wild-type-roots_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 ("Villar_617-1_wild-type-roots_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: Villar_617-1_wild-type-roots_Rep1_ATH1
Organism: Arabidopsis thaliana
Alias: Columbia-0
Stock Code:
Growth Conditions:
SterilisationSeeds were sterilized in 5% NaOCl for 10 minutes and washed 5 times with sterile distilled water.
ProtocolSeeds were sown on MS agar and grown vertically in long-day light conditions in a controlled growth room for 5 days.
Percentage Agrose10
Temperature22 °C average, 22 °C day, 22 °C night
Humidity60 % average, 60 % day, 60 % night
MediumMurashige & Skoog basal salt mixture
Lighting
Developmental Stage:
Growth Stage(Source: Paradigm Genetics)
Tissue: root tips
Additional Organism Information:
Sample Description
Other Information:
ecotype_habitat

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:0.709205
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-ratio1.093959
Spike_AFFX-r2-Ec-bioB_3-5-ratio0.955034
Spike_AFFX-r2-Bs-dap_5_signal97.209846
NoiseAvg:2.62,Std:0.11,Min:2.4,Max:3.0
Control DirectionAntisense
Spike_AFFX-r2-Ec-bioB_5_detectionP
Spike_AFFX-r2-Ec-bioB_3_detectionP
Spike_AFFX-r2-Bs-thr_5_signal25.487267
#P14647
Spike_AFFX-r2-Bs-phe_M_signal66.946030
Corner-Avg:12561,Count:32
Spike_AFFX-r2-P1-cre_3_detectionP
Spike_AFFX-r2-Ec-bioB_M_signal123.512634
Spike_AFFX-r2-Ec-bioB_3_signal91.846443
Spike_AFFX-r2-Bs-lys_M_signal46.360291
Spike_AFFX-r2-P1-cre_3_signal5272.596191
Spike_AFFX-r2-Bs-lys_3-5-ratio2.842206
Spike_AFFX-r2-Bs-dap_M_signal350.320557
Spike_AFFX-r2-Bs-thr_M_detectionP
Probe Pair Threshold1
Spike_AFFX-r2-Bs-thr_3-5-ratio13.046558
Spike_AFFX-r2-Ec-bioB_avg-signal103.843292
Spike_AFFX-r2-Bs-thr_avg-signal170.831482
Spike_AFFX-r2-Bs-thr_5_detectionP
Spike_AFFX-r2-Bs-dap_3_detectionP
Spike_AFFX-r2-Ec-bioD_5_signal925.384766
Spike_AFFX-r2-Bs-phe_5_signal67.620285
Spike_AFFX-r2-P1-cre_5_detectionP
Spike_AFFX-r2-Ec-bioD_3_signal1638.116211
RawQ2.537200
Spike_AFFX-r2-Bs-lys_5_signal24.556870
Signal(A)4.032429
%A34.270058
Signal(All)146.421951
Corner+Avg:118,Count:32
Spike_AFFX-r2-Ec-bioD_5_detectionP
Spike_AFFX-r2-Ec-bioC_3_signal316.245697
Spike_AFFX-r2-Bs-lys_3_detectionP
Spike_AFFX-r2-Bs-phe_3_signal142.584229
Spike_AFFX-r2-Ec-bioD_avg-signal1281.750488
Spike_AFFX-r2-Bs-dap_5_detectionP
Spike_AFFX-r2-Bs-phe_5_detectionP
%P64.213066
Spike_AFFX-r2-Bs-lys_avg-signal46.904278
Spike_AFFX-r2-P1-cre_avg-signal5046.167480
Spike_AFFX-r2-Ec-bioB_M_detectionP
Spike_AFFX-r2-Bs-lys_M_detectionP
#Probe Sets Exceeding Probe Pair Threshold22810
%M1.516879
Spike_AFFX-r2-Bs-thr_3_signal332.521118
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-signal92.383514
Spike_AFFX-r2-Bs-dap_3_signal662.579224
Spike_AFFX-r2-Ec-bioD_3-5-ratio1.770200
Spike_AFFX-r2-Ec-bioD_3_detectionP
Spike_AFFX-r2-Ec-bioC_5_signal333.660339
#M346
Spike_AFFX-r2-Bs-lys_5_detectionP
Spike_AFFX-r2-Bs-lys_3_signal69.795677
Spike_AFFX-r2-Bs-dap_M_detectionP
Spike_AFFX-r2-Ec-bioC_3-5-ratio0.947807
Spike_AFFX-r2-Bs-thr_M_signal154.486053
Signal(P)225.503159
Spike_AFFX-r2-Bs-phe_3-5-ratio2.108601
Spike_AFFX-r2-Bs-thr_3_detectionP
Central-Avg:10059,Count:9
Spike_AFFX-r2-P1-cre_5_signal4819.738770
Spike_AFFX-r2-Bs-phe_M_detectionP
#A7817
Signal(M)15.660192
BackgroundAvg:64.17,Std:0.57,Min:62.2,Max:65.4
Spike_AFFX-r2-Ec-bioC_avg-signal324.953003
Spike_AFFX-r2-Bs-dap_avg-signal370.036530
Spike_AFFX-r2-Bs-dap_3-5-ratio6.815968
Spike_AFFX-r2-Ec-bioB_5_signal96.170815
ScaleMaskAll
BG2
VZ4
TGT100
Alpha10.040000
Alpha20.060000
SF0.709205
NF1.000000
HZ4
Tau0.015000

Slide: Villar_617-2_wild-type-roots_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 ("Villar_617-2_wild-type-roots_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: Villar_617-2_wild-type-roots_Rep2_ATH1
Organism: Arabidopsis thaliana
Alias: Columbia-0
Stock Code:
Growth Conditions:
SterilisationSeeds were sterilized in 5% NaOCl for 10 minutes and washed 5 times with sterile distilled water.
ProtocolSeeds were sown on MS agar and grown vertically in long-day light conditions in a controlled growth room for 5 days.
Percentage Agrose10
Temperature22 °C average, 22 °C day, 22 °C night
Humidity60 % average, 60 % day, 60 % night
MediumMurashige & Skoog basal salt mixture
Lighting
Developmental Stage:
Growth Stage(Source: Paradigm Genetics)
Tissue: root tips
Additional Organism Information:
Sample Description
Other Information:
ecotype_habitat

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:0.729629
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-ratio1.096072
Spike_AFFX-r2-Ec-bioB_3-5-ratio1.091224
Spike_AFFX-r2-Bs-dap_5_signal110.971352
NoiseAvg:2.58,Std:0.07,Min:2.4,Max:2.8
Control DirectionAntisense
Spike_AFFX-r2-Ec-bioB_5_detectionP
Spike_AFFX-r2-Ec-bioB_3_detectionP
Spike_AFFX-r2-Bs-thr_5_signal34.482224
#P14707
Spike_AFFX-r2-Bs-phe_M_signal77.430206
Corner-Avg:12676,Count:32
Spike_AFFX-r2-P1-cre_3_detectionP
Spike_AFFX-r2-Ec-bioB_M_signal119.303169
Spike_AFFX-r2-Ec-bioB_3_signal107.609200
Spike_AFFX-r2-Bs-lys_M_signal60.402321
Spike_AFFX-r2-P1-cre_3_signal5948.318359
Spike_AFFX-r2-Bs-lys_3-5-ratio1.880106
Spike_AFFX-r2-Bs-dap_M_signal380.800415
Spike_AFFX-r2-Bs-thr_M_detectionP
Probe Pair Threshold1
Spike_AFFX-r2-Bs-thr_3-5-ratio10.027029
Spike_AFFX-r2-Ec-bioB_avg-signal108.508545
Spike_AFFX-r2-Bs-thr_avg-signal180.029678
Spike_AFFX-r2-Bs-thr_5_detectionP
Spike_AFFX-r2-Bs-dap_3_detectionP
Spike_AFFX-r2-Ec-bioD_5_signal1210.646484
Spike_AFFX-r2-Bs-phe_5_signal60.300190
Spike_AFFX-r2-P1-cre_5_detectionP
Spike_AFFX-r2-Ec-bioD_3_signal1802.906372
RawQ2.543228
Spike_AFFX-r2-Bs-lys_5_signal34.141258
Signal(A)3.994249
%A34.085926
Signal(All)146.248474
Corner+Avg:104,Count:32
Spike_AFFX-r2-Ec-bioD_5_detectionP
Spike_AFFX-r2-Ec-bioC_3_signal392.555206
Spike_AFFX-r2-Bs-lys_3_detectionP
Spike_AFFX-r2-Bs-phe_3_signal139.931534
Spike_AFFX-r2-Ec-bioD_avg-signal1506.776367
Spike_AFFX-r2-Bs-dap_5_detectionP
Spike_AFFX-r2-Bs-phe_5_detectionP
%P64.476105
Spike_AFFX-r2-Bs-lys_avg-signal52.910931
Spike_AFFX-r2-P1-cre_avg-signal5687.628906
Spike_AFFX-r2-Ec-bioB_M_detectionP
Spike_AFFX-r2-Bs-lys_M_detectionP
#Probe Sets Exceeding Probe Pair Threshold22810
%M1.437966
Spike_AFFX-r2-Bs-thr_3_signal345.754272
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-signal92.553978
Spike_AFFX-r2-Bs-dap_3_signal799.270447
Spike_AFFX-r2-Ec-bioD_3-5-ratio1.489210
Spike_AFFX-r2-Ec-bioD_3_detectionP
Spike_AFFX-r2-Ec-bioC_5_signal376.984680
#M328
Spike_AFFX-r2-Bs-lys_5_detectionP
Spike_AFFX-r2-Bs-lys_3_signal64.189201
Spike_AFFX-r2-Bs-dap_M_detectionP
Spike_AFFX-r2-Ec-bioC_3-5-ratio1.041303
Spike_AFFX-r2-Bs-thr_M_signal159.852570
Signal(P)224.381042
Spike_AFFX-r2-Bs-phe_3-5-ratio2.320582
Spike_AFFX-r2-Bs-thr_3_detectionP
Central-Avg:10440,Count:9
Spike_AFFX-r2-P1-cre_5_signal5426.938965
Spike_AFFX-r2-Bs-phe_M_detectionP
#A7775
Signal(M)14.939817
BackgroundAvg:65.99,Std:0.52,Min:63.4,Max:67.1
Spike_AFFX-r2-Ec-bioC_avg-signal384.769958
Spike_AFFX-r2-Bs-dap_avg-signal430.347382
Spike_AFFX-r2-Bs-dap_3-5-ratio7.202494
Spike_AFFX-r2-Ec-bioB_5_signal98.613258
ScaleMaskAll
BG2
VZ4
TGT100
Alpha10.040000
Alpha20.060000
SF0.729629
NF1.000000
HZ4
Tau0.015000

Slide: Villar_617-3_wild-type-roots_Rep3_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 ("Villar_617-3_wild-type-roots_Rep3_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: Villar_617-3_wild-type-roots_Rep3_ATH1
Organism: Arabidopsis thaliana
Alias: Columbia-0
Stock Code:
Growth Conditions:
SterilisationSeeds were sterilized in 5% NaOCl for 10 minutes and washed 5 times with sterile distilled water.
ProtocolSeeds were sown on MS agar and grown vertically in long-day light conditions in a controlled growth room for 5 days.
Percentage Agrose10
Temperature22 °C average, 22 °C day, 22 °C night
Humidity60 % average, 60 % day, 60 % night
MediumMurashige & Skoog basal salt mixture
Lighting
Developmental Stage:
Growth Stage(Source: Paradigm Genetics)
Tissue: root tips
Additional Organism Information:
Sample Description
Other Information:
ecotype_habitat

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:1.398958
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-ratio1.120723
Spike_AFFX-r2-Ec-bioB_3-5-ratio1.264395
Spike_AFFX-r2-Bs-dap_5_signal91.530571
NoiseAvg:2.07,Std:0.11,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_signal35.297615
#P13388
Spike_AFFX-r2-Bs-phe_M_signal45.102116
Corner-Avg:10403,Count:32
Spike_AFFX-r2-P1-cre_3_detectionP
Spike_AFFX-r2-Ec-bioB_M_signal176.540695
Spike_AFFX-r2-Ec-bioB_3_signal154.713776
Spike_AFFX-r2-Bs-lys_M_signal39.448700
Spike_AFFX-r2-P1-cre_3_signal8190.123047
Spike_AFFX-r2-Bs-lys_3-5-ratio2.245478
Spike_AFFX-r2-Bs-dap_M_signal275.712006
Spike_AFFX-r2-Bs-thr_M_detectionP
Probe Pair Threshold1
Spike_AFFX-r2-Bs-thr_3-5-ratio9.030665
Spike_AFFX-r2-Ec-bioB_avg-signal151.205460
Spike_AFFX-r2-Bs-thr_avg-signal166.600418
Spike_AFFX-r2-Bs-thr_5_detectionP
Spike_AFFX-r2-Bs-dap_3_detectionP
Spike_AFFX-r2-Ec-bioD_5_signal1317.298340
Spike_AFFX-r2-Bs-phe_5_signal40.722164
Spike_AFFX-r2-P1-cre_5_detectionP
Spike_AFFX-r2-Ec-bioD_3_signal2023.971680
RawQ2.239150
Spike_AFFX-r2-Bs-lys_5_signal21.124453
Signal(A)6.823257
%A39.364315
Signal(All)150.702591
Corner+Avg:84,Count:32
Spike_AFFX-r2-Ec-bioD_5_detectionP
Spike_AFFX-r2-Ec-bioC_3_signal509.853058
Spike_AFFX-r2-Bs-lys_3_detectionP
Spike_AFFX-r2-Bs-phe_3_signal121.654312
Spike_AFFX-r2-Ec-bioD_avg-signal1670.635010
Spike_AFFX-r2-Bs-dap_5_detectionP
Spike_AFFX-r2-Bs-phe_5_detectionP
%P58.693554
Spike_AFFX-r2-Bs-lys_avg-signal36.002548
Spike_AFFX-r2-P1-cre_avg-signal7749.007813
Spike_AFFX-r2-Ec-bioB_M_detectionP
Spike_AFFX-r2-Bs-lys_M_detectionP
#Probe Sets Exceeding Probe Pair Threshold22810
%M1.942131
Spike_AFFX-r2-Bs-thr_3_signal318.760956
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-signal69.159531
Spike_AFFX-r2-Bs-dap_3_signal617.248657
Spike_AFFX-r2-Ec-bioD_3-5-ratio1.536457
Spike_AFFX-r2-Ec-bioD_3_detectionP
Spike_AFFX-r2-Ec-bioC_5_signal467.567291
#M443
Spike_AFFX-r2-Bs-lys_5_detectionP
Spike_AFFX-r2-Bs-lys_3_signal47.434494
Spike_AFFX-r2-Bs-dap_M_detectionP
Spike_AFFX-r2-Ec-bioC_3-5-ratio1.090438
Spike_AFFX-r2-Bs-thr_M_signal145.742676
Signal(P)251.414948
Spike_AFFX-r2-Bs-phe_3-5-ratio2.987422
Spike_AFFX-r2-Bs-thr_3_detectionP
Central-Avg:8312,Count:9
Spike_AFFX-r2-P1-cre_5_signal7307.892090
Spike_AFFX-r2-Bs-phe_M_detectionP
#A8979
Signal(M)23.288681
BackgroundAvg:74.45,Std:0.94,Min:72.3,Max:76.5
Spike_AFFX-r2-Ec-bioC_avg-signal488.710175
Spike_AFFX-r2-Bs-dap_avg-signal328.163757
Spike_AFFX-r2-Bs-dap_3-5-ratio6.743634
Spike_AFFX-r2-Ec-bioB_5_signal122.361946
ScaleMaskAll
BG2
VZ4
TGT100
Alpha10.040000
Alpha20.060000
SF1.398958
NF1.000000
HZ4
Tau0.015000

Slide: Villar_617-4_clf-roots_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 ("Villar_617-4_clf-roots_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: Villar_617-4_clf-roots_Rep1_ATH1
Organism: Arabidopsis thaliana
Alias: clf-29
Stock Code:
Genetic Background: Col-0
Growth Conditions:
SterilisationSeeds were sterilized in 5% NaOCl for 10 minutes and washed 5 times with sterile distilled water.
ProtocolSeeds were sown on MS agar and grown vertically in long-day light conditions in a controlled growth room for 5 days.
Percentage Agrose10
Temperature22 °C average, 22 °C day, 22 °C night
Humidity60 % average, 60 % day, 60 % night
MediumMurashige & Skoog basal salt mixture
Lighting
Developmental Stage:
Growth Stage(Source: Paradigm Genetics)
Genetic Variation: T-DNA knock out mutation in AT2G23380
Tissue: root tips
Additional Organism Information:
Sample Description
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:1.245721
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-ratio1.202789
Spike_AFFX-r2-Ec-bioB_3-5-ratio1.034450
Spike_AFFX-r2-Bs-dap_5_signal65.909180
NoiseAvg:2.26,Std:0.06,Min:2.1,Max:2.5
Control DirectionAntisense
Spike_AFFX-r2-Ec-bioB_5_detectionP
Spike_AFFX-r2-Ec-bioB_3_detectionP
Spike_AFFX-r2-Bs-thr_5_signal20.681402
#P13774
Spike_AFFX-r2-Bs-phe_M_signal38.907471
Corner-Avg:10357,Count:32
Spike_AFFX-r2-P1-cre_3_detectionP
Spike_AFFX-r2-Ec-bioB_M_signal133.856491
Spike_AFFX-r2-Ec-bioB_3_signal97.157410
Spike_AFFX-r2-Bs-lys_M_signal24.820251
Spike_AFFX-r2-P1-cre_3_signal6027.482422
Spike_AFFX-r2-Bs-lys_3-5-ratio2.944687
Spike_AFFX-r2-Bs-dap_M_signal226.082947
Spike_AFFX-r2-Bs-thr_M_detectionP
Probe Pair Threshold1
Spike_AFFX-r2-Bs-thr_3-5-ratio10.149895
Spike_AFFX-r2-Ec-bioB_avg-signal108.311890
Spike_AFFX-r2-Bs-thr_avg-signal113.734711
Spike_AFFX-r2-Bs-thr_5_detectionP
Spike_AFFX-r2-Bs-dap_3_detectionP
Spike_AFFX-r2-Ec-bioD_5_signal1055.220215
Spike_AFFX-r2-Bs-phe_5_signal46.489658
Spike_AFFX-r2-P1-cre_5_detectionP
Spike_AFFX-r2-Ec-bioD_3_signal1667.943481
RawQ2.338253
Spike_AFFX-r2-Bs-lys_5_signal19.095091
Signal(A)6.179268
%A37.851818
Signal(All)148.907349
Corner+Avg:80,Count:32
Spike_AFFX-r2-Ec-bioD_5_detectionP
Spike_AFFX-r2-Ec-bioC_3_signal403.388702
Spike_AFFX-r2-Bs-lys_3_detectionP
Spike_AFFX-r2-Bs-phe_3_signal69.664085
Spike_AFFX-r2-Ec-bioD_avg-signal1361.581787
Spike_AFFX-r2-Bs-dap_5_detectionP
Spike_AFFX-r2-Bs-phe_5_detectionP
%P60.385796
Spike_AFFX-r2-Bs-lys_avg-signal33.381466
Spike_AFFX-r2-P1-cre_avg-signal5519.369141
Spike_AFFX-r2-Ec-bioB_M_detectionP
Spike_AFFX-r2-Bs-lys_M_detectionP
#Probe Sets Exceeding Probe Pair Threshold22810
%M1.762385
Spike_AFFX-r2-Bs-thr_3_signal209.914063
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-signal51.687073
Spike_AFFX-r2-Bs-dap_3_signal422.583618
Spike_AFFX-r2-Ec-bioD_3-5-ratio1.580659
Spike_AFFX-r2-Ec-bioD_3_detectionP
Spike_AFFX-r2-Ec-bioC_5_signal342.814178
#M402
Spike_AFFX-r2-Bs-lys_5_detectionP
Spike_AFFX-r2-Bs-lys_3_signal56.229057
Spike_AFFX-r2-Bs-dap_M_detectionP
Spike_AFFX-r2-Ec-bioC_3-5-ratio1.176698
Spike_AFFX-r2-Bs-thr_M_signal110.608665
Signal(P)242.101151
Spike_AFFX-r2-Bs-phe_3-5-ratio1.498486
Spike_AFFX-r2-Bs-thr_3_detectionP
Central-Avg:8515,Count:9
Spike_AFFX-r2-P1-cre_5_signal5011.256348
Spike_AFFX-r2-Bs-phe_M_detectionP
#A8634
Signal(M)21.203203
BackgroundAvg:79.28,Std:0.48,Min:77.7,Max:80.3
Spike_AFFX-r2-Ec-bioC_avg-signal373.101440
Spike_AFFX-r2-Bs-dap_avg-signal238.191910
Spike_AFFX-r2-Bs-dap_3-5-ratio6.411605
Spike_AFFX-r2-Ec-bioB_5_signal93.921776
ScaleMaskAll
BG2
VZ4
TGT100
Alpha10.040000
Alpha20.060000
SF1.245721
NF1.000000
HZ4
Tau0.015000

Slide: Villar_617-5_clf-roots_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 ("Villar_617-5_clf-roots_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: Villar_617-5_clf-roots_Rep2_ATH1
Organism: Arabidopsis thaliana
Alias: clf-29
Stock Code:
Genetic Background: Col-0
Growth Conditions:
SterilisationSeeds were sterilized in 5% NaOCl for 10 minutes and washed 5 times with sterile distilled water.
ProtocolSeeds were sown on MS agar and grown vertically in long-day light conditions in a controlled growth room for 5 days.
Percentage Agrose10
Temperature22 °C average, 22 °C day, 22 °C night
Humidity60 % average, 60 % day, 60 % night
MediumMurashige & Skoog basal salt mixture
Lighting
Developmental Stage:
Growth Stage(Source: Paradigm Genetics)
Genetic Variation: T-DNA knock out mutation in AT2G23380
Tissue: root tips
Additional Organism Information:
Sample Description
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:1.034279
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-ratio1.080521
Spike_AFFX-r2-Ec-bioB_3-5-ratio1.175563
Spike_AFFX-r2-Bs-dap_5_signal133.559494
NoiseAvg:2.36,Std:0.05,Min:2.2,Max:2.5
Control DirectionAntisense
Spike_AFFX-r2-Ec-bioB_5_detectionP
Spike_AFFX-r2-Ec-bioB_3_detectionP
Spike_AFFX-r2-Bs-thr_5_signal35.965904
#P13998
Spike_AFFX-r2-Bs-phe_M_signal88.051964
Corner-Avg:11369,Count:32
Spike_AFFX-r2-P1-cre_3_detectionP
Spike_AFFX-r2-Ec-bioB_M_signal148.573532
Spike_AFFX-r2-Ec-bioB_3_signal126.043953
Spike_AFFX-r2-Bs-lys_M_signal58.317078
Spike_AFFX-r2-P1-cre_3_signal7227.051270
Spike_AFFX-r2-Bs-lys_3-5-ratio2.071684
Spike_AFFX-r2-Bs-dap_M_signal412.050781
Spike_AFFX-r2-Bs-thr_M_detectionP
Probe Pair Threshold1
Spike_AFFX-r2-Bs-thr_3-5-ratio12.906807
Spike_AFFX-r2-Ec-bioB_avg-signal127.279182
Spike_AFFX-r2-Bs-thr_avg-signal228.544083
Spike_AFFX-r2-Bs-thr_5_detectionP
Spike_AFFX-r2-Bs-dap_3_detectionP
Spike_AFFX-r2-Ec-bioD_5_signal1395.235840
Spike_AFFX-r2-Bs-phe_5_signal77.006210
Spike_AFFX-r2-P1-cre_5_detectionP
Spike_AFFX-r2-Ec-bioD_3_signal2056.042969
RawQ2.467519
Spike_AFFX-r2-Bs-lys_5_signal36.851204
Signal(A)5.206502
%A36.931171
Signal(All)151.439606
Corner+Avg:91,Count:32
Spike_AFFX-r2-Ec-bioD_5_detectionP
Spike_AFFX-r2-Ec-bioC_3_signal479.774353
Spike_AFFX-r2-Bs-lys_3_detectionP
Spike_AFFX-r2-Bs-phe_3_signal163.896454
Spike_AFFX-r2-Ec-bioD_avg-signal1725.639404
Spike_AFFX-r2-Bs-dap_5_detectionP
Spike_AFFX-r2-Bs-phe_5_detectionP
%P61.367821
Spike_AFFX-r2-Bs-lys_avg-signal57.170765
Spike_AFFX-r2-P1-cre_avg-signal6957.768066
Spike_AFFX-r2-Ec-bioB_M_detectionP
Spike_AFFX-r2-Bs-lys_M_detectionP
#Probe Sets Exceeding Probe Pair Threshold22810
%M1.701008
Spike_AFFX-r2-Bs-thr_3_signal464.204987
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-signal109.651550
Spike_AFFX-r2-Bs-dap_3_signal913.729431
Spike_AFFX-r2-Ec-bioD_3-5-ratio1.473617
Spike_AFFX-r2-Ec-bioD_3_detectionP
Spike_AFFX-r2-Ec-bioC_5_signal450.950500
#M388
Spike_AFFX-r2-Bs-lys_5_detectionP
Spike_AFFX-r2-Bs-lys_3_signal76.344032
Spike_AFFX-r2-Bs-dap_M_detectionP
Spike_AFFX-r2-Ec-bioC_3-5-ratio1.063918
Spike_AFFX-r2-Bs-thr_M_signal185.461380
Signal(P)243.101028
Spike_AFFX-r2-Bs-phe_3-5-ratio2.128354
Spike_AFFX-r2-Bs-thr_3_detectionP
Central-Avg:8375,Count:9
Spike_AFFX-r2-P1-cre_5_signal6688.484863
Spike_AFFX-r2-Bs-phe_M_detectionP
#A8424
Signal(M)19.457821
BackgroundAvg:82.53,Std:0.67,Min:80.2,Max:83.5
Spike_AFFX-r2-Ec-bioC_avg-signal465.362427
Spike_AFFX-r2-Bs-dap_avg-signal486.446564
Spike_AFFX-r2-Bs-dap_3-5-ratio6.841366
Spike_AFFX-r2-Ec-bioB_5_signal107.220055
ScaleMaskAll
BG2
VZ4
TGT100
Alpha10.040000
Alpha20.060000
SF1.034279
NF1.000000
HZ4
Tau0.015000

Slide: Villar_617-6_clf-roots_Rep3_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 ("Villar_617-6_clf-roots_Rep3_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: Villar_617-6_clf-roots_Rep3_ATH1
Organism: Arabidopsis thaliana
Alias: clf-29
Stock Code:
Genetic Background: Col-0
Growth Conditions:
SterilisationSeeds were sterilized in 5% NaOCl for 10 minutes and washed 5 times with sterile distilled water.
ProtocolSeeds were sown on MS agar and grown vertically in long-day light conditions in a controlled growth room for 5 days.
Percentage Agrose10
Temperature22 °C average, 22 °C day, 22 °C night
Humidity60 % average, 60 % day, 60 % night
MediumMurashige & Skoog basal salt mixture
Lighting
Developmental Stage:
Growth Stage(Source: Paradigm Genetics)
Genetic Variation: T-DNA knock out mutation in AT2G23380
Tissue: root tips
Additional Organism Information:
Sample Description
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:1.16163
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-ratio1.108681
Spike_AFFX-r2-Ec-bioB_3-5-ratio1.030432
Spike_AFFX-r2-Bs-dap_5_signal91.515900
NoiseAvg:2.43,Std:0.09,Min:2.3,Max:2.8
Control DirectionAntisense
Spike_AFFX-r2-Ec-bioB_5_detectionP
Spike_AFFX-r2-Ec-bioB_3_detectionP
Spike_AFFX-r2-Bs-thr_5_signal26.434216
#P13795
Spike_AFFX-r2-Bs-phe_M_signal57.070992
Corner-Avg:11047,Count:32
Spike_AFFX-r2-P1-cre_3_detectionP
Spike_AFFX-r2-Ec-bioB_M_signal145.367813
Spike_AFFX-r2-Ec-bioB_3_signal102.329155
Spike_AFFX-r2-Bs-lys_M_signal39.072662
Spike_AFFX-r2-P1-cre_3_signal6202.166016
Spike_AFFX-r2-Bs-lys_3-5-ratio2.026894
Spike_AFFX-r2-Bs-dap_M_signal319.425110
Spike_AFFX-r2-Bs-thr_M_detectionP
Probe Pair Threshold1
Spike_AFFX-r2-Bs-thr_3-5-ratio12.959020
Spike_AFFX-r2-Ec-bioB_avg-signal115.668007
Spike_AFFX-r2-Bs-thr_avg-signal175.205078
Spike_AFFX-r2-Bs-thr_5_detectionP
Spike_AFFX-r2-Bs-dap_3_detectionP
Spike_AFFX-r2-Ec-bioD_5_signal994.418457
Spike_AFFX-r2-Bs-phe_5_signal69.542564
Spike_AFFX-r2-P1-cre_5_detectionP
Spike_AFFX-r2-Ec-bioD_3_signal1812.905396
RawQ2.442143
Spike_AFFX-r2-Bs-lys_5_signal36.390236
Signal(A)6.299163
%A37.689610
Signal(All)151.178635
Corner+Avg:87,Count:32
Spike_AFFX-r2-Ec-bioD_5_detectionP
Spike_AFFX-r2-Ec-bioC_3_signal383.493469
Spike_AFFX-r2-Bs-lys_3_detectionP
Spike_AFFX-r2-Bs-phe_3_signal125.450661
Spike_AFFX-r2-Ec-bioD_avg-signal1403.661865
Spike_AFFX-r2-Bs-dap_5_detectionP
Spike_AFFX-r2-Bs-phe_5_detectionP
%P60.477859
Spike_AFFX-r2-Bs-lys_avg-signal49.740688
Spike_AFFX-r2-P1-cre_avg-signal5898.174316
Spike_AFFX-r2-Ec-bioB_M_detectionP
Spike_AFFX-r2-Bs-lys_M_detectionP
#Probe Sets Exceeding Probe Pair Threshold22810
%M1.832530
Spike_AFFX-r2-Bs-thr_3_signal342.561523
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-signal84.021400
Spike_AFFX-r2-Bs-dap_3_signal613.956787
Spike_AFFX-r2-Ec-bioD_3-5-ratio1.823081
Spike_AFFX-r2-Ec-bioD_3_detectionP
Spike_AFFX-r2-Ec-bioC_5_signal402.330383
#M418
Spike_AFFX-r2-Bs-lys_5_detectionP
Spike_AFFX-r2-Bs-lys_3_signal73.759163
Spike_AFFX-r2-Bs-dap_M_detectionP
Spike_AFFX-r2-Ec-bioC_3-5-ratio0.953180
Spike_AFFX-r2-Bs-thr_M_signal156.619492
Signal(P)245.399063
Spike_AFFX-r2-Bs-phe_3-5-ratio1.803941
Spike_AFFX-r2-Bs-thr_3_detectionP
Central-Avg:9122,Count:9
Spike_AFFX-r2-P1-cre_5_signal5594.182617
Spike_AFFX-r2-Bs-phe_M_detectionP
#A8597
Signal(M)21.413126
BackgroundAvg:82.30,Std:0.58,Min:80.2,Max:84.4
Spike_AFFX-r2-Ec-bioC_avg-signal392.911926
Spike_AFFX-r2-Bs-dap_avg-signal341.632599
Spike_AFFX-r2-Bs-dap_3-5-ratio6.708745
Spike_AFFX-r2-Ec-bioB_5_signal99.307053
ScaleMaskAll
BG2
VZ4
TGT100
Alpha10.040000
Alpha20.060000
SF1.161630
NF1.000000
HZ4
Tau0.015000


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