GSM6265678: liver, 3e11 AAV8-CSDE1 shRNA, chow diet, rep 3; Mus musculus; RNA-Seq
Samples were collected in Invitrogen TRIzol reagent and RNA was purified using standard protocols. 1 ug of RNA was used for cDNA library construction NEBNext® Ultra Directional RNA Library Prep Kit for Illumina® (#E7420S) according to the manufacturer's protocol. Briefly, mRNA was enriched using oligo(dT) beads followed by two rounds of purification and randomly fragmented by addition of fragmentation buffer. The first strand cDNA was synthesized using random hexamer primers, after which a second-strand synthesis buffer, dNTPs, RNase H, and DNA polymerase I were added to generate the second-strand. After terminal repair and sequencing adaptor ligation, the double-stranded cDNA library underwent size selection and PCR enrichment. The resulting 250-350 bp insert libraries were quantified using a Qubit 2.0 fluorometer and quantitative PCR. Size distribution was analyzed using an Agilent 2100 Bioanalyzer.