Open AccessEvaluation of protocols for rRNA depletion-based RNA sequencing of nanogram inputs of mammalian total RNA
Author(s) -
Simon Haile,
Richard Corbett,
Steve Bilobram,
Karen Mungall,
Bruno M. Grande,
Heather Kirk,
Pawan Pandoh,
Tina MacLeod,
Helen McDonald,
Miruna Bala,
Robin Coope,
Richard A. Moore,
Yongjun Zhao,
Ryan D. Morin,
Steve Jones,
Marco A. Marra
Publication year - 2019
Publication title -
plos one
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.99
H-Index - 332
ISSN - 1932-6203
DOI - 10.1371/journal.pone.0224578
Subject(s) - rna , ribosomal rna , biology , rna splicing , rna editing , computational biology , gene expression , rna extraction , microbiology and biotechnology , gene , genetics
Next generation RNA-sequencing (RNA-seq) is a flexible approach that can be applied to a range of applications including global quantification of transcript expression, the characterization of RNA structure such as splicing patterns and profiling of expressed mutations. Many RNA-seq protocols require up to microgram levels of total RNA input amounts to generate high quality data, and thus remain impractical for the limited starting material amounts typically obtained from rare cell populations, such as those from early developmental stages or from laser micro-dissected clinical samples. Here, we present an assessment of the contemporary ribosomal RNA depletion-based protocols, and identify those that are suitable for inputs as low as 1–10 ng of intact total RNA and 100–500 ng of partially degraded RNA from formalin-fixed paraffin-embedded tissues.