RNAseq data analysis

Quality control was provided by the National Genomics Infrastructure (NGI) at Science for Life Laboratory, Stockholm, Sweden. The first step after mRNA-sequencing was quantifying abundances of transcript sequences in FASTA format by generating abundance estimates for all samples using the Kallisto software [61]. Also, gene abundance estimates were performed by summing the transcript expression (TPM) values for the transcripts of the same gene. Since DESeq2 expects count data, from the Kallisto output the ‘tximport’ package was used to convert these estimates into read counts. DESeq2 was performed as a basis for differential gene expression analysis based on the negative binomial distribution [62]. Low gene counts (<100) were filtered out and variance stabilizing transformation was performed on the count data, as well as batch correction using the ‘limma’ package.