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Virology. 2014 Jan 5;448:168-75. doi: 10.1016/j.virol.2013.10.008. Epub 2013 Oct 25.

Deamination intensity profiling of human APOBEC3 protein activity along the near full-length genomes of HIV-1 and MoMLV by HyperHRM analysis.

Author information

1
Department of Biochemistry, Microbiology and Immunology, Faculty of Medicine, University of Ottawa, Ottawa, Ontario, Canada.

Abstract

Enzymatic deamination of cytidines in DNA is an intrinsic component of antibody maturation and retroviral resistance, but can also be a source of HIV drug resistance and cancer-causing mutations. Here, we developed a high-throughput method based on high resolution melt (HRM) analysis called HyperHRM that can screen genomic DNA for rare hypermutated proviral sequences and accurately quantify the number of C-to-T or G-to-A mutations in each sequence. We demonstrate the effectiveness of the approach by profiling in parallel the intensity of the DNA mutator activity of all seven human APOBEC3 proteins on the near full-length sequence of HIV-1 and the Moloney murine leukemia virus. Additionally, HRM was successfully used to identify hypermutated proviral sequences in peripheral blood mononuclear cells from an HIV-1 patient. These results exemplify the effectiveness of HRM-based approaches for hypermutation quantification and for the detection of hypermutated DNA sequences potentially associated with disease or retroviral drug resistance.

KEYWORDS:

APOBEC3; Cytidine deamination; HIV; Hypermutation; Moloney murine leukemia virus

PMID:
24314647
DOI:
10.1016/j.virol.2013.10.008
[Indexed for MEDLINE]
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