Computational protocol: Optimized approach for Ion Proton RNA sequencing reveals details of RNA splicing and editing features of the transcriptome

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Protocol publication

[…] RNA libraries were sequenced on P1v2 chips using the Ion Proton™ System (Thermo Fisher Scientific, #4476610). Sequencing was completed by the Analytical and Translational Genomics Shared Resource at the University of New Mexico Cancer Center. All samples were aligned with Torrent Mapping Alignment Program (TMAP), using the 1000 genomes GRCh37 phase II d5 the human reference genome. Subsequently, the data were aligned with the STAR aligner [] using the same reference genome. Exon feature counts were generated with HTSeq-count [] using a modified RefSeq references, or our own annotations when counting MYB features. Consistent with the Ion Proton sequencing technology, all reads were treated as single-end for the purposes of genome alignment and feature counting. [...] Cufflinks [] with the Ensembl 75 reference was used to estimate isoform observations []. Subsequent analysis of the data was conducted in R using packages listed in supplemental section 1. RNA sequencing data are available for download from the NCBI Sequence Read Archive using SRA BioProject accession number PRJNA368701. […]

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