The total RNA was extracted with HiPure Universa miRNA Kit (Magen) from two pairs of fresh frozen tissue samples and quality was confirmed by Nanodrop measurement of OD 260/280 and 260/230 ratios. The material for library construction was 1 μg per sample. Sequencing libraries were constructed following the Illumina TruSeq Stranded protocol. Total RNA Gold kit with Ribo-Zero Gold (Illumina, USA) was used following the manufacturer’s recommendations. Sequencing (2×150 paired-end reads) was performed at Mingma Technologies Co., Ltd in Shanghai.
FASTQ format data were assessed using FastQC (v0.11.9) and then fastp (v0.20.1) (35 (link)) was used to remove the bases with an average quality value less than 20 and to cut the reads of adapters. Clean reads were mapped to the human reference genome (Gencode v19) by STAR (2.7.5b) (36 (link)). The gene and transcript isoform expression was quantified using RSEM (v1.3.1) (37 (link)). Bedtools (v2.29.2) (38 (link)) was used to transform the bam files to bw format for UCSC genome browser viewing.
FASTQ format data were assessed using FastQC (v0.11.9) and then fastp (v0.20.1) (35 (link)) was used to remove the bases with an average quality value less than 20 and to cut the reads of adapters. Clean reads were mapped to the human reference genome (Gencode v19) by STAR (2.7.5b) (36 (link)). The gene and transcript isoform expression was quantified using RSEM (v1.3.1) (37 (link)). Bedtools (v2.29.2) (38 (link)) was used to transform the bam files to bw format for UCSC genome browser viewing.
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