Transgenic Arabdopsis thaliana 35S::CKX1 seedlings overexpressing cytokinin oxidase grown in liquid MS were treated for 15 minutes with 6-Benzyl adenine dissolved in diluted KOH. Total RNA was extracted with the TRIzol method using the high salt precipitation described in the troubleshooting section of the TRIzol manual. The RNA was further purified using Qiagen mini columns. 5 µg of the purified total RNA was used for cDNA synthesis using Invitrogen SuperScriptII reverse transcriptase and a mixture of Ligase, DNA polymerase, and RNase and purified using Qiagen PCR purification columns. The whole amount of cDNA was used as a template to synthesize Biotin-labeled cRNA using the Enzo High Yield RNA transcript labeling kit. After fragmentation as recommended by Affymetrix, the sample was submitted to the RZPD (Berlin), where the subsequent steps were performed to generate a table containing the normalized expression values and expression calls.
