Title |
Duplex-specific nuclease efficiently removes rRNA for prokaryotic RNA-seq
|
---|---|
Published in |
Nucleic Acids Research, August 2011
|
DOI | 10.1093/nar/gkr617 |
Pubmed ID | |
Authors |
Hana Yi, Yong-Joon Cho, Sungho Won, Jong-Eun Lee, Hyung Jin Yu, Sujin Kim, Gary P. Schroth, Shujun Luo, Jongsik Chun |
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. |
X Demographics
Geographical breakdown
Country | Count | As % |
---|---|---|
Unknown | 2 | 100% |
Demographic breakdown
Type | Count | As % |
---|---|---|
Members of the public | 2 | 100% |
Mendeley readers
Geographical breakdown
Country | Count | As % |
---|---|---|
United States | 9 | 4% |
Italy | 3 | 1% |
Sweden | 2 | <1% |
United Kingdom | 2 | <1% |
Estonia | 2 | <1% |
Australia | 1 | <1% |
Germany | 1 | <1% |
Canada | 1 | <1% |
New Zealand | 1 | <1% |
Other | 6 | 2% |
Unknown | 224 | 89% |
Demographic breakdown
Readers by professional status | Count | As % |
---|---|---|
Researcher | 81 | 32% |
Student > Ph. D. Student | 56 | 22% |
Student > Master | 20 | 8% |
Professor > Associate Professor | 13 | 5% |
Student > Postgraduate | 11 | 4% |
Other | 45 | 18% |
Unknown | 26 | 10% |
Readers by discipline | Count | As % |
---|---|---|
Agricultural and Biological Sciences | 151 | 60% |
Biochemistry, Genetics and Molecular Biology | 40 | 16% |
Environmental Science | 9 | 4% |
Immunology and Microbiology | 7 | 3% |
Engineering | 3 | 1% |
Other | 12 | 5% |
Unknown | 30 | 12% |