For each time point (T0, T1 and T2) one leaflet each of 14, 6 and 9 genotypes in genotypic groups A1, A2 and B2, respectively, were pooled. Frozen pooled leaf tissue was powdered in a CryoMill (Retsch, Haan, Germany). Total RNA was extracted from powdered tissue using the Plant RNA Isolation Kit of Invitrogen (Karlsruhe, Germany) following the manufacturer’s protocol. RNA purification, removal of genomic DNA contamination, concentration measurements and quality tests were performed as described (Draffehn et al. 2010). RNA integrity was additionally assessed by electrophoresis using an Agilent 2100 Bioanalyzer (Agilent Technologies, Santa Clara, USA). SuperSAGE libraries were generated at GenXPro GmbH (Frankfurt, Germany) essentially as described (Matsumura et al. 2010). To prevent amplification biases the TrueQuant technology was applied as described by B. Rotter (Patent application Nr. WO2009152928). Besides NlaIII (recognition site: 5’-CATG-3’), DpnII (recognition site: 5’-GATC-3’) was used as second anchoring enzyme, to capture transcripts without a NlaIII site. Therefore, the 26 bp tags carry either CATG or GATC at their 5’ end. The libraries were pooled and sequenced by Solexa/Illumina technology (Illumina, Inc., USA).
GEO Accession
GSM1167249