title: GSM7674862: Scramble Knockdown total bio rep 1; Homo sapiens; RNA-Seq
library_construction_protocol: For RNA isolation, purification was carried out using a mixture of phenol/chloroform/isoamyl alcohol (25:24:1, pH 6.7) (Fisher, BP1752I-100) and vortexed for 30 sec. The samples were then centrifuged at 14,000 rpm for 10 min at 4C. The top aqueous phase was transferred to a new tube containing 3 M sodium acetate, 2 µg of glycogen and 250 µl of absolute ethanol, and was then left for overnight precipitation at -80C. The resulting pellet was washed with prechilled 70% ethanol before centrifuging at the same speed. DNase I (NEB, Cat #M0303L) was added to the RNA pellet and incubated at 37oC for 10 min, followed by treatment with 50 mM EDTA (Bioworld, Cat #40520000-1) to eliminate DNA contamination. The RNA pellet was then purified as described above one more time to obtain pure RNA samples. RNA libraries for RNA-seq were prepared using NGS Library Prep Kit following manufacturer's protocols.