title: GSM8756138: MDA-MB-231, CTR, RNA-seq, replicate 3; Homo sapiens; RNA-Seq
library_construction_protocol: MCF7, T47D, MDA-MB-231 and Hs578T TIS cells were harvested at day 12, HFF TIS cells were harvested at day 24. CTR and REPOP cells were harvested at ~80% confluency. Cells were homogenized in TRIzol™ Reagent (Thermo Fisher Scientific). Total RNA was extracted from samples using Direct-zol® MiniPrep kit (Zymo Research) following the manufacturer's instructions. To prevent DNA contamination, an in-column DNAse I treatment was performed.  The RNA concentration was determined by using the Qubit RNA HS Assay Kit on the Qubit 3.0 Fluorometer (Thermo Fisher Scientific). Quality CTR was assessed by Labchip GX Touch HT instrument on DNA 5K/RNA/CZE Chip (Perkin Elmer) with RNA Pico Sensitivity Assay Reagents (Perkin Elmer). NextFlex PolyA beads 2.0 kit and NextFlex Rapid Directional RNA-seq Kit 2.0 with UDIs (Perkin Elmer) were used for mRNA capture and strand-specific library preparation. The library quantities were measured by Quant-iT 1x dsDNA HS Assay kit (Thermo Fisher Scientific) with Fluostar Omega (BMG Labtech). The fragment size distribution of the libraries was determined by capillary electrophoresis on Labchip GX Touch Nucleic Acid Analyzer on XMark HT chip by using DNA NGS 3k Assay kit (Perkin Elmer). Pooled libraries were sequenced with 50M 150 bp paired-end reads on NovaSeq 6000 platform (Illumina).