Transcriptome Sequencing of cold-stress Jatropha curcas L.
Tissue samples from three cold treatments and the control seedlings of Jatropha curcas L., were equally mixed for RNA preparation. Total RNA was extracted using a TRIzol reagent (Invitrogen) following the manufacture’s protocol. The concentration of the RNA was determined with Qubit Fluorometer, and the quality was checked by Agilent 2100 Bioanalyzer and 1% (w/v) agarose gel electrophoresis. By using oligo(dT)-attached magnetic beads, poly(A)-containing mRNAs were enriched from the total RNA extracts. Fragmentation buffer was added for interrupting mRNA to short segments, which were taken as the templates to synthesize cDNAs by random hexamer-primers. Then, the short DNA products were purified with QiaQuick PCR extraction kit and ligated with sequencing adapters. By agarose gel electrophoresis, suitable fragments with 200-700bp were selected for PCR amplification and sequencing templates by the Illumina Hiseq™ 2000 RNA-seq system.