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Article: A qRT-PCR-based method for the measurement of rrn operon copy number

TitleA qRT-PCR-based method for the measurement of rrn operon copy number
Authors
Issue Date2009
Citation
Letters In Applied Microbiology, 2009, v. 49 n. 1, p. 26-30 How to Cite?
AbstractAim: To develop a convenient and accurate method for estimating the rrn operon copy number (Yrrn) in cells of pure prokaryotic cultures based on quantitative real-time polymerase chain reaction (qRT-PCR). Methods & Results: Using Escherichia coli, the Yrrn of which is known to be 7, as a reference, the rrn concentrations of target species and E. coli in sample solutions were measured based on their respective threshold cycle numbers (Ct), whereas the cell concentrations of both species were measured by microscopic counting after staining. The Yrrn of the target species was then calculated from the initial cell concentrations and the rrn concentrations of the target species and E. coli. Using this method, the Y rrn values of four species, i.e. Xanthomonas campestris, Staphylococcus aureus, Aeromonas hydrophila and Pseudomonas fluorescens, were estimated as 1·80, 4·73, 8·58 and 5·13, respectively, comparable to their respective known values of 2, 5, 10, and 5, resulting in an average deviation of 8%. Conclusions: The whole cell qRT-PCR based methods were convenient, accurate and reproducible in quantification of rrn copy number of prokaryotic cells. Significance and Impact of the Study: qTR-PCR is a fast and reliable DNA quantification approach. Compared with previous qTR-PCR based methods measuring rrn copy number, the present method avoided the prerequisite for the information on genome size and GC content of target bacteria or a gene with known copy number, thus should be more widely applicable. © 2009 The Society for Applied Microbiology.
Persistent Identifierhttp://hdl.handle.net/10722/150496
ISSN
2015 Impact Factor: 1.579
2015 SCImago Journal Rankings: 0.641
ISI Accession Number ID
Funding AgencyGrant Number
Hong Kong Research Grants CouncilHKU 7195/06E
Funding Information:

The authors wish to thank the Hong Kong Research Grants Council for the financial support of this study (HKU 7195/06E), and Ming-Fei Shao wishes to thank HKU for the postgraduate studentship.

References
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DC FieldValueLanguage
dc.contributor.authorZhang, Ten_US
dc.contributor.authorShao, MFen_US
dc.contributor.authorFang, HHPen_US
dc.date.accessioned2012-06-26T06:05:10Z-
dc.date.available2012-06-26T06:05:10Z-
dc.date.issued2009en_US
dc.identifier.citationLetters In Applied Microbiology, 2009, v. 49 n. 1, p. 26-30en_US
dc.identifier.issn0266-8254en_US
dc.identifier.urihttp://hdl.handle.net/10722/150496-
dc.description.abstractAim: To develop a convenient and accurate method for estimating the rrn operon copy number (Yrrn) in cells of pure prokaryotic cultures based on quantitative real-time polymerase chain reaction (qRT-PCR). Methods & Results: Using Escherichia coli, the Yrrn of which is known to be 7, as a reference, the rrn concentrations of target species and E. coli in sample solutions were measured based on their respective threshold cycle numbers (Ct), whereas the cell concentrations of both species were measured by microscopic counting after staining. The Yrrn of the target species was then calculated from the initial cell concentrations and the rrn concentrations of the target species and E. coli. Using this method, the Y rrn values of four species, i.e. Xanthomonas campestris, Staphylococcus aureus, Aeromonas hydrophila and Pseudomonas fluorescens, were estimated as 1·80, 4·73, 8·58 and 5·13, respectively, comparable to their respective known values of 2, 5, 10, and 5, resulting in an average deviation of 8%. Conclusions: The whole cell qRT-PCR based methods were convenient, accurate and reproducible in quantification of rrn copy number of prokaryotic cells. Significance and Impact of the Study: qTR-PCR is a fast and reliable DNA quantification approach. Compared with previous qTR-PCR based methods measuring rrn copy number, the present method avoided the prerequisite for the information on genome size and GC content of target bacteria or a gene with known copy number, thus should be more widely applicable. © 2009 The Society for Applied Microbiology.en_US
dc.languageengen_US
dc.relation.ispartofLetters in Applied Microbiologyen_US
dc.subject.meshBacteria - Geneticsen_US
dc.subject.meshGene Dosageen_US
dc.subject.meshGenes, Rrnaen_US
dc.subject.meshOperonen_US
dc.subject.meshRna, Bacterial - Geneticsen_US
dc.subject.meshReverse Transcriptase Polymerase Chain Reaction - Methodsen_US
dc.titleA qRT-PCR-based method for the measurement of rrn operon copy numberen_US
dc.typeArticleen_US
dc.identifier.emailZhang, T:zhangt@hkucc.hku.hken_US
dc.identifier.emailFang, HHP:hrechef@hkucc.hku.hken_US
dc.identifier.authorityZhang, T=rp00211en_US
dc.identifier.authorityFang, HHP=rp00115en_US
dc.description.naturelink_to_subscribed_fulltexten_US
dc.identifier.doi10.1111/j.1472-765X.2009.02613.xen_US
dc.identifier.pmid19413774-
dc.identifier.scopuseid_2-s2.0-67249139272en_US
dc.identifier.hkuros161880-
dc.relation.referenceshttp://www.scopus.com/mlt/select.url?eid=2-s2.0-67249139272&selection=ref&src=s&origin=recordpageen_US
dc.identifier.volume49en_US
dc.identifier.issue1en_US
dc.identifier.spage26en_US
dc.identifier.epage30en_US
dc.identifier.isiWOS:000266924300005-
dc.publisher.placeUnited Kingdomen_US
dc.relation.projectMicrobial diversity and characteristics of activated sludge treating Hong Kong's saline sewage-
dc.identifier.scopusauthoridZhang, T=24470677400en_US
dc.identifier.scopusauthoridShao, MF=34868583400en_US
dc.identifier.scopusauthoridFang, HHP=7402542625en_US
dc.identifier.citeulike4863562-

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