Samples of frozen tumor tissues were first homogenized using a MagNA Lyser device (Roche, Basel, Switzerland). Subsequent total RNA isolation was performed using the MagNA Pure Compact RNA Kit (Roche) on the MagNA Pure Compact System (Roche) according to the manufacturer’s protocol. The concentration of isolated total RNA was assessed on a Qubit 4.0 fluorimeter (Thermo Fisher Scientific, Waltham, MA, USA) using the Qubit RNA BR Assay Kit (Thermo Fisher Scientific). The RIN (RNA integrity number) parameter, which characterizes the integrity of RNA, was evaluated using an Agilent 2100 Bioanalyzer (Agilent Technologies, Santa Clara, CA, USA). The RIN for all samples studied was no less than 7.
Sample preparation of mRNA libraries was performed using the TruSeq Stranded mRNA Kit (Illumina, San Diego, CA, USA) as described previously [35 (link)]. The size of the resulting mRNA library was ~260 bp.
High-throughput sequencing of mRNA libraries was performed on a NextSeq 500 System (Illumina) using NextSeq 500/550 High Output Kit v2.5 (Illumina) in 75 bp single-ended read mode. On average, about 20 million reads were received for each sample.
Sample preparation of mRNA libraries was performed using the TruSeq Stranded mRNA Kit (Illumina, San Diego, CA, USA) as described previously [35 (link)]. The size of the resulting mRNA library was ~260 bp.
High-throughput sequencing of mRNA libraries was performed on a NextSeq 500 System (Illumina) using NextSeq 500/550 High Output Kit v2.5 (Illumina) in 75 bp single-ended read mode. On average, about 20 million reads were received for each sample.
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