GSM9006067: RNAseq, PANC-1 cell, shSTYK1-1; Homo sapiens; RNA-Seq
Total RNA was extracted using Trizol, and an Oligo(dT) magnetic beads  were used to isolate mRNA. ThismRNA was fragmented and synthesized into a one-stranded cDNA using random primers for reverse synthesis, followed by two-stranded synthesis to form a stable duplex structure. After end repair, ligation of adapters, fragment screening and library enrichment, followed by on-board sequencing. mRNA was isolated according to polyA selection method by oligo(dT) beads and then fragmented by fragmentation buffer firstly. Secondly double-stranded cDNA was synthesized using a SuperScript double-stranded cDNA synthesis kit (Invitrogen, CA) with random hexamer primers (Illumina). Then the synthesized cDNA was subjected to end-repair, phosphorylation and 'A' base addition according to Illumina's library construction protocol. Libraries were size selected for cDNA target fragments of 300 bp on 2% Low Range Ultra Agarose followed by PCR amplified using Phusion DNA polymerase (NEB) for 15 PCR cycles. After quantified by Qubit 4.0, paired-end RNA-seq sequencing library was sequenced with the NovaSeq X plus sequencer (2 × 150bp read length).