Detection and analysis of DNA methylation by pyrosequencing.

Hilary A. A. Colyer, Richard N. Armstrong, Daniel J. Sharpe, Ken I. Mills

Research output: Contribution to journalBook/Film/Article review

6 Citations (Scopus)

Abstract

Pyrosequencing is a “sequencing by synthesis” technique which can be used to quantify DNA methylation at specific CpG sites within the target region of interest. Biotin labelled polymerase chain reaction (PCR) products form the template for base-pair nucleotide incorporation causing a light emitting cascade reaction resulting in the formation of a pyrogram and the calculation of the percentage methylation for each site. Prior to pyrosequencing, it is essential to bisulphite-convert the DNA sample and then perform locus-specific PCR for the region of interest. One of the PCR primers needs to be biotinylated and a separate sequencing primer is required for the pyrosequencing itself.
Original languageEnglish
Pages (from-to)281-292
Number of pages12
JournalMethods in Molecular Biology
Volume863
Early online date03 Feb 2012
DOIs
Publication statusPublished - 2012

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DNA Methylation
Polymerase Chain Reaction
Biotin
Base Pairing
Methylation
Nucleotides
Light
DNA

Cite this

Colyer, Hilary A. A. ; Armstrong, Richard N. ; Sharpe, Daniel J. ; Mills, Ken I. / Detection and analysis of DNA methylation by pyrosequencing. In: Methods in Molecular Biology . 2012 ; Vol. 863. pp. 281-292.
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Detection and analysis of DNA methylation by pyrosequencing. / Colyer, Hilary A. A.; Armstrong, Richard N.; Sharpe, Daniel J.; Mills, Ken I.

In: Methods in Molecular Biology , Vol. 863, 2012, p. 281-292.

Research output: Contribution to journalBook/Film/Article review

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