GSM8292087: K562, WT, rep3; Homo sapiens; RNA-Seq
Total RNA was isolated using TRIzol (Invitrogen) reagent according to the manufacturer's protocol. mRNA was enriched from total RNA using oligo(dT) beads. First, the mRNA was randomly fragmented by adding fragmentation buffer. Then, cDNA was synthesized using the mRNA as a template and random hexamers primer. A custom second-strand synthesis buffer (Illumina), dNTPs, RNase H, and DNA polymerase I were added to initiate the second-strand synthesis. After a series of terminal repairs, A-tailing, and sequencing adaptor ligation, the double-stranded cDNA library was completed through size selection and PCR enrichment.