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PLoS One. 2019 Oct 31;14(10):e0224578. doi: 10.1371/journal.pone.0224578. eCollection 2019.

Evaluation of protocols for rRNA depletion-based RNA sequencing of nanogram inputs of mammalian total RNA.

Author information

1
Canada's Michael Smith Genome Sciences Centre, BC Cancer, Vancouver, British Columbia, Canada.
2
Department of Molecular Biology and Biochemistry, Simon Fraser University, Burnaby, British Columbia, Canada.
3
Department of Medical Genetics, University of British Columbia, Vancouver, British Columbia, Canada.

Abstract

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.

Conflict of interest statement

The authors have declared that no competing interests exist.

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