title: GSM6604356: Huh-7 cells-2; Homo sapiens; RNA-Seq
library_construction_protocol: Total RNA was isolated using RNeasy mini kit (Qiagen, Germany). Total RNA was quantified using a NanoDrop ND-2000 spectrophotometer (Thermo Fisher Scientific, Waltham, MA, USA) The total RNA samples are first treated with DNase I to degrade any possible DNA contamination. Then the mRNA is enriched by using the oligo(dT) magnetic beads (foreukaryotes). Mixed with the fragmentation buffer, the mRNA is fragmented into short fragments (about 200 bp). Then the first strand of cDNA is synthesized by using random.  hexamer-primer. Buffer, dNTPs, RNase H and DNA polymerase I are added to synthesize the second strand. The double strand cDNA is purified with magnetic beads. End  reparation and 3'-end single nucleotide A (adenine) addition is then performed. Finally, sequencing adaptors are ligated to the fragments. The fragments are enriched by PCR  amplification. During the QC step, Agilent 2100 Bioanaylzer and ABI StepOnePlus Real-Time PCR System are used to qualify and quantify of the sample library.