RNA-seq was performed as by MARS-seq as described in [55 (link)]. The bulk MARS-seq libraries were sequenced with high-output 75-bp kits (FC-404-2005, Illumina, USA) on NextSeq 500/550 Illumina sequencer.
Processing of raw sequencing data into read counts was performed via the User-friendly Transcriptome Analysis Pipeline (UTAP) [56 (link)]. Reads were trimmed using cutadapt [57 ] and mapped to genome (/shareDB/iGenomes/Mus_musculus/UCSC/mm10/Sequence/STAR_index) using STAR [58 (link)] (default parameters). The pipeline quantifies genes annotated in RefSeq (that have expanded with 1,000 bases toward 5′ edge and 100 bases toward 3′ bases). Counting (UMI counts) was done using HTSeq-count in union mode [59 (link)]. Count normalization was performed using DESeq2 [60 (link)] with the following parameters: betaPrior = True, cooksCutoff = FALSE, and independentFiltering = FALSE.
RNA-seq data are available from the GEO database (accession number GSE171975). All other data that support the findings of this study are available from the corresponding author upon request.
Processing of raw sequencing data into read counts was performed via the User-friendly Transcriptome Analysis Pipeline (UTAP) [56 (link)]. Reads were trimmed using cutadapt [57 ] and mapped to genome (/shareDB/iGenomes/Mus_musculus/UCSC/mm10/Sequence/STAR_index) using STAR [58 (link)] (default parameters). The pipeline quantifies genes annotated in RefSeq (that have expanded with 1,000 bases toward 5′ edge and 100 bases toward 3′ bases). Counting (UMI counts) was done using HTSeq-count in union mode [59 (link)]. Count normalization was performed using DESeq2 [60 (link)] with the following parameters: betaPrior = True, cooksCutoff = FALSE, and independentFiltering = FALSE.
RNA-seq data are available from the GEO database (accession number GSE171975). All other data that support the findings of this study are available from the corresponding author upon request.
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