NASC's International Affymetrix Service

AI logo

NASCArrays

home, selection, spot history, gene swinger, two gene scatterplot, digital northern, bulk gene download, tutorial, help, Affymetrix site, NASC

Experiment: NPA-induced vascular overgrowth in Arabidopsis

Experiment Description

NASCArrays Experiment Reference Number: NASCARRAYS-428

We used NPA-induced vascular overgrowth in Arabidopsis leaves to look for differential up-regulation of genes in NPA-treated tissues that may be involved in vascular differentiation. Arabidopsis thaliana Col-0 plants were grown for approximately 2 weeks on solid ATS medium (1) containing a final concentration of 10 um NPA (dissolved in DMSO) or an equivalent volume of DMSO (control). At this stage plants had approximately 6 rosette leaves. RNA was prepared from entire shoot tissues of control (DMSO) or NPA-treated plants.(1) Lincoln et al., 1990. Plant Cell 2: 1071-1080.

About the Experimenter

Name:Dr Carol Wenzel
Head of Lab Name:Dr Jim Mattsson
Lab:
Address:Department of Biological Sciences
Simon Fraser University
8888 University Drive
Burnaby, BC
Postcode: V5A 1S6
Country: Canada
 
Telephone Number: 17787824391
Fax Number: 17787823496

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:2
 
Experimental Parameters:
parametercompound_based_treatment
Quality Control Measures Taken:
References:
 
Other Information:
GEO AccessionGSE19262

Slides in this Experiment

Hybridisation Set: Wenzel: NPA-induced vascular overgrowth in Arabidopsis_genome

Slide: Wenzel_1-1_DMSO-control-2wk-shoots_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 ("Wenzel_1-1_DMSO-control-2wk-shoots_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: Wenzel_1-1_DMSO-control-2wk-shoots_Rep1_ATH1
Organism: Arabidopsis thaliana
Alias: Col-0
Stock Code: N1092
Growth Conditions:
stratification4C for at least 2 days
sterilisation3 hr chlorine gas
Growth protocolSeeds of Arabidopsis thaliana ecotype Col-0 were sterilized with chlorine gas and plated onto sterile ATS media (Lincoln et al., 1990) supplemented with either N-(1-Naphthyl) phthalamic acid (NPA; Sigma, St. Louis, MO) dissolved in DMSO to a final concentration of 10 mM, or an equivalent volume of DMSO for control material. Seeds were stratified for at least 2 days at 4°C and then grown at ca 20°C with 16h illumination for 2 weeks (shoot material). Shoot tissues were immediately immersed in liquid nitrogen after excision, stored at -80°C, and total RNA purified according to Chang et al. (1993). Lincoln et al. 1990. Plant Cell 2: 1071-1080 Chang et al. 1993. Biology Reporter 11: 113-116
LocationGrowth Room
Growth substrateAgar
Average humidity70%
Average temperature20oC
Growth mediumOwn medium: see Lincoln et al. 1990. Plant Cell 2: 1071-1080
Sample descriptionshoot tissue from ca 2wk old plants with ca 6 rosette leaves
Developmental Stage:
Developmental stage (Boyes Key)1.04
Tissue: shoot
Other Information:

Protocols for BioSource 1

Extraction Protocol Information

Name of Protocol: Change method
Method:
Protocolsee Chang et al. 1993. Biology Reporter 11: 113-116
Type: total RNA
Other Information:
ProtocolSourceSubmitter

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:0.566161
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:
RawQ2.27
NoiseAvg:3.41,Stdev:0.17,Max:3.9,Min:2.9
BackgroundAvg:60.92,Stdev:0.90,Max:64.0,Min:58.9
Gamma1H0.0025
Gamma2L0.003
Gamma2H0.003
TGT100
Perturbation1.1
SF0.566161
Alpha10.04
Alpha20.06
SFGeneAll
NF1.000000
BF
Gamma1L0.0025
Tau0.015

Slide: Wenzel_1-2_NPA-treated-2wk-shoots_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 ("Wenzel_1-2_NPA-treated-2wk-shoots_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: Wenzel_1-2_NPA-treated-2wk-shoots_Rep1_ATH1
Organism: Arabidopsis thaliana
Alias: Col-0
Stock Code: N1092
Growth Conditions:
stratification4C for at least 2 days
sterilisation3 hr chlorine gas
stratification4C for at least 2 days
sterilisation3 hr chlorine gas
Growth protocolSeeds of Arabidopsis thaliana ecotype Col-0 were sterilized with chlorine gas and plated onto sterile ATS media (Lincoln et al., 1990) supplemented with either N-(1-Naphthyl) phthalamic acid (NPA; Sigma, St. Louis, MO) dissolved in DMSO to a final concentration of 10 mM, or an equivalent volume of DMSO for control material. Seeds were stratified for at least 2 days at 4°C and then grown at ca 20°C with 16h illumination for 2 weeks (shoot material). Shoot tissues were immediately immersed in liquid nitrogen after excision, stored at -80°C, and total RNA purified according to Chang et al. (1993). Lincoln et al. 1990. Plant Cell 2: 1071-1080 Chang et al. 1993. Biology Reporter 11: 113-116
LocationGrowth Room
Growth substrateAgar
Average humidity70%
Average temperature20oC
Growth mediumOwn medium: see Lincoln et al. 1990. Plant Cell 2: 1071-1080. Modification: add N-(1-Naphthyl) phthalamic acid (NPA; Sigma, St. Louis, MO) dissolved in DMSO to a final concentration of 10 uM
Sample descriptionshoot tissue from ca 2wk old plants with ca 6 rosette leaves
Developmental Stage:
Developmental stage (Boyes Key)1.04
Tissue: shoot
in vivo Treatment:
treatmentadded N-(1-Naphthyl) phthalamic acid (NPA; Sigma, St. Louis, MO) dissolved in DMSO to a final concentration of 10 uM
Other Information:

Protocols for BioSource 1

Extraction Protocol Information

Name of Protocol: Change method
Method:
Protocolsee Chang et al. 1993. Biology Reporter 11: 113-116
Type: total RNA
Other Information:
ProtocolSourceSubmitter

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:0.467506
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:
RawQ2.24
NoiseAvg:3.26,Stdev:0.16,Max:3.9,Min:2.9
BackgroundAvg:62.43,Stdev:0.97,Max:64.9,Min:60.0
Gamma1H0.0025
Gamma2L0.003
Gamma2H0.003
TGT100
Perturbation1.1
SF0.467506
Alpha10.04
Alpha20.06
SFGeneAll
NF1.000000
BF
Gamma1L0.0025
Tau0.015


Problems? Comments? Suggestions? Contact the Affymetrix Team