GSM3389628: HEK-293T_siHNRNPH1_rep1; Homo sapiens; RNA-Seq
RNA was extracted with Maxwell® 16 LEV simplyRNA Cells Kit. RNA quantity was measured using the Nanodrop. Poly(A)-selected RNA from three biological replicates and sequencing libraries were created using TruSeq Stranded mRNA LT Sample Preparation Kit as described by manufacturer's instructions (Illumina, San Diego, CA). Then, we incorporated a capture step using fosmid clones of full genomic coverage for EWSR1 (WIBR2-3436F12; GRCh37/hg19 chr22:29662533-29698996) or EWS-FLI1 (WIBR2-3436F12 and WIBR2-1520I3; GRCh37/hg19 chr11:128640224-128682737). Capture bait was prepared by Klenow extension of biotinylated random hexamers priming on purified fosmid DNA template. The prepared bait was bound to streptavidin-coated Dynabeads C1 (Invitrogen) and hybridized to denatured, pooled libraries for 48 hours. The enriched library was recovered after a series of stringency washes by PCR using primers homologous to library end-sequences (Forward: 5'-AATGATACGGCGACCACC-3', Reverse: 5'-CAAGCAGAAGACGGCATA-3'). Libraries were quantified by qPCR (Kapa Biosystems, Wilmington, MA).