title: GSM6923463: RNA-seq_hAFE_rep2; Homo sapiens; RNA-Seq
library_construction_protocol: Total RNA was isolated using Trizol Reagent (Invitrogen, 15596-018) according to the manufacturer's instructions. For quantitative PCR analysis, 1 μg of total RNA was reverse-transcribed using SuperScript III First-Strand Synthesis System (Thermo, 18080051). Bulk RNA-sequencing library preparation was performed following a published protocol {Zhang:2012kc}. Briefly, 4 μg of total RNA was depleted of ribosomal RNA using Ribo-Zero rRNA Removal Kit (Epicentre, MRZH116) and then converted to cDNA. Final sequencing libraries were generated using dUTP (Thermo, R0133) incorporation and uracil-N-glycosylase (NEB, M0280) treatment to generate strand-specific RNA-seq libraries. The final libraries were sequenced at paired-end on a Hi-Seq2000 platform.