E-MEXP-2858 - Transcription profiling by array of Arabidopsis wild type and transgenic lines grown under normal or salt stress conditions
Submitted on 9 August 2010, last updated on 3 June 2014, released on 30 July 2014
Arabidopsis ATH1 Genome Array was used to study the expression levels of 22,500 probe-sets representing 24,000 genes. Both wild type and different transgenic lines were allowed to grow for one month under normal condition was taken for expression analysis. These plants were treated with half strength of Hoagland solution supplemented with 200mM NaCl in pots for seven days for salinity treatment. High quality RNA was extracted from the healthy leaf samples using TRI Reagent (Ambion, INC. USA) and pooled from at least four independent stressed and non-stressed samples separately, and treated with DNase-I (QIAGEN GmbH, Germany). RNA cleanup was carried out using RNeasy Plant Mini Kit (QIAGEN GmbH, Germany) and 5µg of total RNA from each sample with three biological replications were reverse-transcribed to double stranded cDNA using the GeneChipR One-Cycle cDNA Synthesis Kit. The biotin-labelled cRNA was made using the GeneChipR IVT Labelling Kit (Affymetrix, CA, USA). Twenty microgram of cRNA samples was fragmented and was hybridized for 16 hours at 45°C to the Affymetrix Arabidopsis ATH1 Genome Array (Santa Clara, CA, USA). After washing and staining with R-phycoerythrin streptavidin in a Fluidics Station, using the GenechipR Fluidics Station 450, the arrays were scanned by the GenechipR 3000 Scanner. The chip images were scanned and extracted using default settings and the .CEL files were generated using Affymetrix GeneChip Command Console (AGCC) Software.
Atleast two ATH1 Genome Array was used for hybridisation for each group of samples.
transcription profiling by array, co-expression, compound treatment, replicate