GSM1645628: 93-11_CT; Oryza sativa; RNA-Seq
Total RNA was extracted with the mirVana miRNA Isolation Kit (Life Technologies) from Nipponbare and 93-11 seedlings with and without chilling treatment. Sequencing libraries were generated using NEBNext® Ultra™ RNA Library Prep Kit for Illumina® (NEB) following manufacturer’s recommendations. Briefly, The mRNAs were enriched using oligo(dT) magnetic beads and broken into short fragments. First strand cDNA was synthesized using random hexamer primer and M-MuLV Reverse Transcriptase. Second strand cDNA synthesis was subsequently performed using DNA Polymerase I. NEBNext Adaptor was ligated. The fragments were purified with Agencourt AMPure XP Kit (Beckman Coulter) to select cDNA fragments of preferentially 150~200 bp. PCR amplification was performed with size-selected and adaptor-ligated cDNA with Phusion High-Fidelity DNA Polymerase. PCR products were purified and sequenced on an Illumina Hiseq 2000 platform.