Labelled using protocol described in Affymetrix manual "3' IVT Express Kit User Manual"
Hybridization protocol
Biotin Labelled cRNA using "Affymetrix GeneChip Hybridization, Wash and Stain Kit", according to guidelines set out by Affymetrix, Santa Clara, CA
Scan protocol
Scanned according to guidelines set out by Affymetrix, Santa Clara, CA.
Description
2 days, at 4 degrees. 20% bleach solution for 10 minutes, washed 3 x with distilled water. Arabidopsis thaliana seeds were sterilized in 20% bleach for 10 min and washed with sterilized water for 3 times. Seeds were sown in Petri-dishes containing sterilized media, Murashige and Skoog (MS) basal salt pH 5.7, 1% sucrose and 0.5% phytagel (Sigma, UK). Prior to germination, seeds were stratified at 4oC for 4 days. The plants were grown vertically in the growth room with continual light at 25oC. Autoclaved 25oC average, 25oC day, 25oC night on plates % average, on plates % day, on plates % night Murashige & Skoog basal salt mixture. Modifications: Murashige and Skoog (MS) basal salt pH 5.7, 1% sucrose and 0.5% phytagel (Sigma, UK) Constant light
Data processing
Raw data from the microarrays was normalized at probe-level using MAS5 algorithm. The detection calls (present, marginal, absent) for each probe set was obtained using the GCOS system
