GSM9104596: H1703 cells, SAHA, Repeat3; Homo sapiens; RNA-Seq
Total RNA was extracted from cells grown in 6-well plates (2 × 105 cells per well, ~70 % confluence at harvest) using TRIzol reagent (Invitrogen) and purified with the RNeasy Mini Kit (Qiagen), including an on-column DNase I digestion. RNA purity (A260/280 ratio = 2.05 ± 0.04, A260/230 ratio = 1.98 ± 0.03) was measured using NanoDrop 2000, and RNA integrity (RIN ≥ 9.6) was confirmed by an Agilent 2100 Bioanalyzer. High-quality RNA (500 ng per sample) was used to generate strand-specific poly(A) libraries with the NEBNext Ultra II Directional RNA Library Prep Kit. Poly-(A)-tailed mRNA was isolated from total RNA extracted from human cell lines and converted to cDNA with random hexamer priming. A non-directional (un-stranded) Illumina library was prepared according to the manufacturer's standard protocol (comparable to TruSeq RNA v2), without rRNA depletion. Adapters with unique dual indices were ligated, fragments were size-selected (~300 bp insert), PCR-amplified, quantified, pooled, and sequenced on an Illumina NovaSeq 6000 (PE150) platform.