title: GSM8640719: RNA-Seq, epithelium, control, rep 3; Homo; RNA-Seq
library_construction_protocol: RNA was extracted and purified using TRIzol (Thermo Fisher, 15596018). A concentration>50ng/µL, RIN value>7.0, and total RNA>1µg are sufficient for downstream experiments. Use oligo (dT) magnetic beads (Dynabeads Oligo (dT), cat. 25-61005, Thermo Fisher, USA) to specifically capture mRNA containing PolyA (polyadenylation) through two rounds of purification. The captured mRNA was fragmented using the NEBNextR Magnesium RNA Fragmentation Module (cat. E6150S, USA) under high temperature conditions at 94 ℃ for 5-7 minutes. cDNA was synthesized using Invitrogen SuperScriptTM II Reverse Transcriptase (cat. 1896649, CA, USA), and E Escherichia coli DNA polymerase I (NEB, cat. m0209, USA) and RNase H (NEB, cat. m0297, USA) were subjected to double stranded synthesis. The double stranded DNA was digested using UDG enzyme (NEB, cat. m0280, MA, US), and a library with fragment size of 300bp ± 50bp was formed by PCR (chain specific library)