GSM6255089: siCTR_Rep2; Homo sapiens; RNA-Seq
Total RNA was isolated from MCF-7 cells using the RNeasy Mini Kit, according to the manufacturer's protocol (Qiagen, 74104, Hilden, Germany). Experiments were performed in three biological replicates. First-strand cDNA synthesis was performed with the SensiFAST™ cDNA Synthesis Kit (Bioline, BIO-65054, London, UK). Qualitative PCR analysis was performed using MA038 EconoTaq 2xMaster Mix (Lucigen). PCR products were analyzed on a 3% agarose gel, stained with SYBR™ Safe DNA Gel Stain (S33102 ThermoFisher Scientific). RNA quality check (RNA integrity number (RIN) > 8) was achieved with a Fragment Analyzer (Advanced Analytical Technologies, Inc., Ankeny, IA, USA) and quantified with Qubit (Qubit™ RNA HS Assay Kit, ThermoFisher Scientific, Waltham, MA, USA; Q32852). Total RNA, depleted of ribosomal RNAs fractions, was used to construct the RNA-Seq libraries using Illumina TruSeq RNA sample prep kit (TruSeq™ RNA Sample Prep Kit v2-Set B, Illumina, RS-122-2002, San Diego, CA, USA).