QuantSeq 3' mRNA-Seq Library Preparation

The QuantSeq 3’ mRNA-Seq Library Prep Kit FWD for Illumina (Lexogen Inc, Greenland, NH, USA) and the UMI Second Strand Synthesis Module for QuantSeq FWD (Illumina Inc, San Diego, CA, USA) were used to construct a sequencing library. The libraries were sequenced on a NextSeq device (Illumina Inc, San Diego, CA, USA) as a single-end 76. The unique molecular identifiers were removed from the raw reads with UMI-tools (v1.1.2). The quality and the length of the raw reads were inspected utilizing FastQC (v0.11.9) [19 ]. FastQ Screen (v0.15.0) [20 (link)] and a set of common lab organism genomes were used to check for putative contaminations. Up to 95% of the sequenced reads map on the bovine genome, whereas the non-bovine reads did not map to the tested lab organisms. The quality of the reads based on phred scores was good. Cutadapt (v3.7) [21 ] was used to trim the adaptors and filter reads containing ambiguities or reads that did not pass the phred score threshold of 20. Finally, the quality was checked with FastQc (v0.11.9) [19 ].