Open AccessncRNAseq: simple modifications to RNA-seq library preparation allow recovery and analysis of mid-sized non-coding RNAs
Author(s) -
Nicola Minshall,
Igor Chernukhin,
Jason S. Carroll,
Anna Git
Publication year - 2022
Publication title -
biotechniques/biotechniques
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.617
H-Index - 131
eISSN - 1940-9818
pISSN - 0736-6205
DOI - 10.2144/btn-2021-0035
Subject(s) - computational biology , rna , coding (social sciences) , genome , biology , deep sequencing , long non coding rna , computer science , genetics , gene , mathematics , statistics
Despite their abundance, mid-sized RNAs (30–300 nt) have not been extensively studied by high-throughput sequencing, mostly due to selective loss in library preparation. The authors propose simple and inexpensive modifications to the Illumina TruSeq protocol (ncRNAseq), allowing the capture and sequencing of mid-sized non-coding RNAs without detriment to the coverage of coding mRNAs. This protocol is coupled with a two-step alignment: a pre-alignment to a curated non-coding genome, passing only the non-mapping reads to a standard genomic alignment. ncRNAseq correctly assigns the highest read-numbers to established abundant non-coding RNAs and correctly identifies cytosolic and nuclear enrichment of known non-coding RNAs in two cell lines.