Quantitation of site-specific HPV 16 DNA methylation by pyrosequencing.

Quantitation of site-specific HPV 16 DNA methylation by pyrosequencing. Research Abstract Details 

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Quantitation of site-specific HPV 16 DNA methylation by pyrosequencing. Abstract Text:

Mangalathu S Rajeevan,David C Swan,Kara Duncan,Daisy R Lee,Josef R Limor,Elizabeth R Unger,

Human papillomavirus (HPV) is a necessary but insufficient cause of cervical cancer. Factors influencing transcription, such as epigenetic silencing through viral DNA methylation, may impact neoplastic progression. Pyrosequencing technology was applied to quantify methylation at 19 cytosine guanine dinucleotide (CpG) sites in the L1 3' and long control region (LCR) of HPV 16 DNA using cell lines, CaSki ( approximately 400 integrated copies of HPV 16) and SiHa (1-2 integrated copies of HPV 16) that differ in their transcriptional activity. Methylation levels ranged from 20 to 100% in CaSki and from 0 to 85% in SiHa over the entire 19 CpG sites, with a >40-fold difference in the methylation levels of their promoter and enhancer regions (SiHa<2% and CaSki 79%). The method was successful at a limiting dilution of 1-4 HPV 16 DNA copies/3000 cells, a level compatible with most clinical samples. The results were not affected by fixation in methanol-based liquid cytology collection fluid or method of extraction. Conditions optimized with cell lines were applicable to fixed exfoliated cervical cells. Pyrosequencing provides a quantitative site-specific assessment of methylation at multiple CpG sites without cloning, and is thus suited to large-scale molecular epidemiologic studies.

Quantitation of site-specific HPV 16 DNA methylation by pyrosequencing. Publishing Authors By Initials

MS Rajeevan,DC Swan,K Duncan,DR Lee,JR Limor,ER Unger,

For similar diagnosis: laboratory techniques and procedures: specimen handling research abstracts see: diagnosis: laboratory techniques and procedures: specimen handling research

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Quantitation of site-specific HPV 16 DNA methylation by pyrosequencing. Journal Published:

PUBLICATION TYPE: Research Support, U.S. Gov't,

Journal: Journal of virological methods

VOLUME: 138

Page Numbers: 170-6

Journal Abbreviation: J. Virol. Methods

ISSN: 0166-0934

DAY: 11

MONTH: 10

YEAR: 2006

Quantitation of site-specific HPV 16 DNA methylation by pyrosequencing. Information

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LANGUAGE: eng

NlmUniqueID: 8005839

Quantitation of site-specific HPV 16 DNA methylation by pyrosequencing. Keywords Mesh Terms:

KEYWORDS: Specimen Handling

MESH TERMS: methods

Chemical & Substance for Abstract: Quantitation of site-specific HPV 16 DNA methylation by pyrosequencing. Information

Substance Name: DNA, Viral

Registry Number: 0

Grant and Affiliation Information for Quantitation of site-specific HPV 16 DNA methylation by pyrosequencing.

AFFILIATION: Division of Viral and Rickettsial Diseases, National Center for Infectious Diseases, Centers for Disease Control & Prevention, 1600 Clifton Road, MSG41, Atlanta, GA 30333, USA. mor4@cdc.gov

Country: Netherlands

AGENCY: United States NCI

GRANT: Y1-CN-5005-01

ACRONYM: CN

MEDLINETA: J Virol Methods

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