GSM1693042: UIP rep2; Sus scrofa; RNA-Seq
Total RNA was extracted from longissimus dorsi using TRIzol (Invitrogen, CA, USA) and further purified with RNeasy column (Qiagen, USA) according to the manufacturer’s protocol Approximately 10 ug of total RNA representing a muscle tissue was sub-jected to isolate Poly (A) mRNA with poly-T oligo attached magnetic beads (Thermo-fisher). Following purification, the mRNA was fragmented into small pieces using divalent cations under elevated temperature. Then the cleaved RNA fragments were constructed into the final cDNA library in accordance with the protocol for the Illumina RNA ligation based method (Illumina, San Diego, USA). In brief, the fragmented RNA has been dephosphorylated at the 3' end by the phosphatase and been phosphorylated at the 5' end by the PNK. The treated RNA has been cleaned up with the RNeasy MinElute Kit (Qiagen) following the instructions of the manufacturer. The purified RNA was ligated with a pre-adenylated 3' adapter, which enables the subsequent ligation of the 5' adapter. Based on the adapter sequence, a reverse transcription followed by PCR used to create cDNA constructs. The average insert size for the single-end libraries was 300 bp (±50 bp). And then we performed the single end sequencing on an Illumina Hiseq2000 at the (LC Sciences,USA) following the vendor's recommended protocol.
GEO Accession
GSM1693042