RNA-Seq of monoclonal cell lines after CRISPR/Cas9-mediated editing of the small nucleolar RNA host-gene GAS5

In this study monoclonal cell lines carrying mutations in the small nucleolar RNA host-gene GAS5 were obtained. Next, Poly(A) RNA-seq and Small RNA-seq analyses of obtained cell lines were performed on an Illumina NextSeq 500 platform. Poly(A) RNA-seq libraries was constructed on polyA mRNA-enriched fraction. Small RNA-seq libraries was constructed on small RNA fraction (<200 nucleotide length). The reads were aligned via STAR to the human genome (GRCh37). The comparison of the mRNA levels of genes and small RNA levels in RNA-Seq experiments was carried out using CuffDiff tool. Sashimi plots for RNA-Seq analyses of isoform expression were generated by IGV. It was shown that specific mutations in SNORD74 led to the downregulation of all GAS5-encoded SNORDs and GAS5 lncRNA. Obtained results demonstrate the SNORD-dependent manner of the GAS5 maturation.