GSM9825615: HUVEC, LPS, OE-vec-2; Homo sapiens; RNA-Seq
Total RNA was extracted from cultured HUVEC cells using TRIzol reagent (Invitrogen). RNA purity was detected by NanoPhotometer spectrophotometer, RNA concentration was quantified by Qubit 2.0 Fluorometer, and RNA integrity was assessed by Agilent 2100 Bioanalyzer. Samples with RIN value higher than 7.0 were used for subsequent library construction. Genomic DNA contamination was removed via DNase I digestion. mRNA was enriched from total RNA by oligo(dT) magnetic beads, then fragmented at 94 degrees C. First-strand cDNA was synthesized with random hexamer primers, followed by second-strand cDNA synthesis. End repair, 3 prime A-tailing and adapter ligation were performed sequentially. Purified cDNA fragments were amplified by PCR to construct strand-specific RNA-seq libraries. Library quality was verified with Agilent DNA 1000 Assay Kit, and qualified libraries were sequenced on Illumina platform with paired-end 150 bp mode.