RNA-Seq of homo sapiens: squamous carcinoma
RNA libraries were generated and sequenced according to Suntsova et al (2019). RNA was extracted using RNeasy Micro Kit (Qiagen). RNA concentrations were measured fluorimetrically with Qubit 4 and Qubit RNA BR Assay Kit (Thermo Fisher Scientific). Tapestation 4150 with RNA ScreenTapes and Reagents (Agilent) were used to measure RNA Integrity Number (RIN). Ribosomal RNA depletion and library construction was performed with KAPA RNA HyperPrep Kit with RiboErase (HMR) (Roche). KAPA unique dual-indexed primers (Roche) were used for multiplexing samples in a single sequencing run. Library concentrations were measured using the Qubit 4 with Qubit ds DNA HS Assay kit (Thermo Fisher Scientific). For fragment length distributions we used Tapestation 4150 with D1000 ScreenTapes and Reagents (Agilent). The RNA sequencing was performed at the Laboratory of Clinical and Genomic Bioinformatics, I.M. Sechenov First Moscow State Medical University (Sechenov University), Moscow, Russia, on Illumina NextSeq 550 equipment for single end sequencing, 75 bp read length, for approximately 20 million raw reads per sample. Data quality check was conducted using Illumina SAV. De-multiplexing was performed using Illumina Bcl2fastq2 v 2.17 software.