期刊
METHODS
卷 63, 期 2, 页码 126-134出版社
ACADEMIC PRESS INC ELSEVIER SCIENCE
DOI: 10.1016/j.ymeth.2013.03.023
关键词
RNA-seq; Transcriptome; Duplex-specific nuclease; Gene expression
资金
- National Cancer Institute [CA085289]
- Integrative Cancer Biology Program [CA1113001, P50 CA083639, U54 CA151668, CA125806]
- Ovarian Cancer Research Fund [PPDIU01.2011]
- National Science Foundation [ABO-1062432]
This report describes an improved protocol to generate stranded, barcoded RNA-seq libraries to capture the whole transcriptome. By optimizing the use of duplex specific nuclease (DSN) to remove ribosomal RNA reads from stranded barcoded libraries, we demonstrate improved efficiency of multiplexed next generation sequencing (NGS). This approach detects expression profiles of all RNA types, including miRNA (microRNA), piRNA (Piwi-interacting RNA), snoRNA (small nucleolar RNA), lincRNA (long non-coding RNA), mtRNA (mitochondrial RNA) and mRNA (messenger RNA) without the use of gel electrophoresis. The improved protocol generates high quality data that can be used to identify differential expression in known and novel coding and non-coding transcripts, splice variants, mitochondrial genes and SNPs (single nucleotide polymorphisms). (C) 2013 Elsevier Inc. All rights reserved.
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