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Duplex-specific nuclease efficiently removes rRNA for prokaryotic RNA-seq

Cited 67 time in Web of Science Cited 77 time in Scopus
Authors
Yi, Hana; Cho, Yong-Joon; Won, Sungho; Lee, Jong-Eun; Yu, Hyung Jin; Kim, Sujin; Schroth, Gary P.; Luo, Shujun; Chun, Jongsik
Issue Date
2011-11
Citation
Nucleic Acids Research, Vol.39 No.20, p. e140
Abstract
Next-generation sequencing has great potential for application in bacterial transcriptomics. However, unlike eukaryotes, bacteria have no clear mechanism to select mRNAs over rRNAs; therefore, rRNA removal is a critical step in sequencing-based transcriptomics. Duplex-specific nuclease (DSN) is an enzyme that, at high temperatures, degrades duplex DNA in preference to single-stranded DNA. DSN treatment has been successfully used to normalize the relative transcript abundance in mRNA-enriched cDNA libraries from eukaryotic organisms. In this study, we demonstrate the utility of this method to remove rRNA from prokaryotic total RNA. We evaluated the efficacy of DSN to remove rRNA by comparing it with the conventional subtractive hybridization (Hyb) method. Illumina deep sequencing was performed to obtain transcriptomes from Escherichia coli grown under four growth conditions. The results clearly showed that our DSN treatment was more efficient at removing rRNA than the Hyb method was, while preserving the original relative abundance of mRNA species in bacterial cells. Therefore, we propose that, for bacterial mRNA-seq experiments, DSN treatment should be preferred to Hyb-based methods.
ISSN
0305-1048
URI
https://hdl.handle.net/10371/165905
DOI
https://doi.org/10.1093/nar/gkr617
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College of Natural Sciences (자연과학대학)Dept. of Biological Sciences (생명과학부)Journal Papers (저널논문_생명과학부)
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