GSM1646810: 24h_RNA; Citrus trifoliata; RNA-Seq
Total RNA was extracted with RNAiso Plus (TaKaRa, Dalian, China) according to the manufacturer's instructions. The cDNA library was constructed using mRNA-Seq assay according to the instructions. In brief, poly(A) mRNA was enriched from 20 μg of total RNA using oligo(dT) magnetic beads, and cut into short fragments (200-700 bp) with fragmentation buffer. The short mRNA fragments were synthesized into double-strand cDNA using random hexamer primers, and purified with QiaQuick PCR Purification Kit (Qiagen, CA, USA). Then cDNA was washed with EB buffer for end repairing and poly(A) addition. Illumina paired-end sequencing adapters were then ligated to the ends of the 3’-adenylated cDNA fragments. Agarose gel electrophoresis was carried out so as to select fragments with suitable size that was enriched by PCR amplification to construct the cDNA library. The library was sequenced by BGI (Shenzhen, China) on Illumina HiSeqTM 2000 platform for 5 gigabase in-depth sequencing.