GSM8636180: NC_1; Homo sapiens; RNA-Seq
Total RNA was treated with RQ1 DNase (Promega) to remove DNA. The quality and quantity of the purified RNA were determined by measuring the absorbance at 260nm/280nm (A260/A280) using smartspec plus (BioRad). RNA integrity was further verified by 1.5% agarose gel electrophoresis. For each sample, 1μg of total RNA was used for RNA-seq library preparation with NEBNext® Ultra™ Directional RNA Library Prep Kit for Illumina® (#E7420). Polyadenylated mRNAs were purified and fragmented, and then converted into double strand cDNA. Following end repair and A tailing, the DNAs were ligated to Diluted NEBNext® Adaptor. After purification, ligation products corresponding to 300-500bps were digested with Heat-labile UDG, the single strand cDNA was amplified, purified, quantified, and stored at -80 ℃ before sequencing. For high-throughput sequencing, the libraries were prepared following the manufacturer's instructions and applied to Illumina NovaSeq 6000 system for 150 nt paired-end sequencing.