Specific rolling circle amplification of low-copy human polyomaviruses BKV, HPyV6, HPyV7, TSPyV, and STLPyV

Rockett, Rebecca, Barraclough, Katherine A., Isbel, Nicole M., Dudley, Kevin J., Nissen, Michael D., Sloots, Theo P., & Bialasiewicz, Seweryn (2015) Specific rolling circle amplification of low-copy human polyomaviruses BKV, HPyV6, HPyV7, TSPyV, and STLPyV. Journal of Virological Methods, 215-216, pp. 17-21.

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Abstract

Eleven new human polyomaviruses have been recently discovered, yet for most of these viruses, little is known of their biology and clinical impact. Rolling circle amplification (RCA) is an ideal method for the amplification of the circular polyomavirus genome due to its high fidelity amplification of circular DNA. In this study, a modified RCA method was developed to selectively amplify a range of polyomavirus genomes. Initial evaluation showed a multiplexed temperature-graded reaction profile gave the best yield and sensitivity in amplifying BK polyomavirus in a background of human DNA, with up to 1 × 10(8)-fold increases in viral genomes from as little as 10 genome copies per reaction. Furthermore, the method proved to be more sensitive and provided a 200-fold greater yield than that of random hexamers based standard RCA. Application of the method to other novel human polyomaviruses showed successful amplification of TSPyV, HPyV6, HPyV7, and STLPyV from low-viral load positive clinical samples, with viral genome enrichment ranging from 1 × 10(8) up to 1 × 10(10). This directed RCA method can be applied to selectively amplify other low-copy polyomaviral genomes from a background of competing non-specific DNA, and is a useful tool in further research into the rapidly expanding Polyomaviridae family.

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ID Code: 90189
Item Type: Journal Article
Refereed: Yes
Keywords: Polyomavirus; Rolling circle amplification; Specific; Clinical sample; Low copy; BKV
DOI: 10.1016/j.jviromet.2015.02.004
ISSN: 0166-0934
Divisions: Current > Schools > School of Earth, Environmental & Biological Sciences
Current > Institutes > Institute for Future Environments
Current > QUT Faculties and Divisions > Science & Engineering Faculty
Copyright Owner: Copyright 2015 Elsevier B.V.
Deposited On: 13 Nov 2015 05:19
Last Modified: 17 Nov 2015 03:07

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