Correction of PCR-bias in quantitative DNA methylation studies by means of cubic polynomial regression

Evgeny A. Moskalev, Mikhail G. Zavgorodnij, Svetlana P. Majorova, Ivan A. Vorobjev, Pouria Jandaghi, Irina V. Bure, Jörg D. Hoheisel

Research output: Contribution to journalArticle

47 Citations (Scopus)

Abstract

DNA methylation profiling has become an important aspect of biomedical molecular analysis. Polymerase chain reaction (PCR) amplification of bisulphite-treated DNA is a processing step that is common to many currently used methods of quantitative methylation analysis. Preferential amplification of unmethylated alleles - known as PCR-bias - may significantly affect the accuracy of quantification. To date, no universal experimental approach has been reported to overcome the problem. This study presents an effective method of correcting biased methylation data. The procedure includes a calibration performed in parallel to the analysis of the samples under investigation. DNA samples with defined degrees of methylation are analysed. The observed deviation of the experimental results from the expected values is used for calculating a regression curve. The equation of the best-fitting curve is then used for correction of the data obtained from the samples of interest. The process can be applied irrespective of the locus interrogated and the number of sites analysed, avoiding an optimization of the amplification conditions for each individual locus.

Original languageEnglish
JournalNucleic Acids Research
Volume39
Issue number11
DOIs
Publication statusPublished - Jun 2011
Externally publishedYes

Fingerprint

DNA Methylation
Methylation
Polymerase Chain Reaction
DNA Fingerprinting
DNA
Calibration
Alleles

ASJC Scopus subject areas

  • Genetics

Cite this

Moskalev, E. A., Zavgorodnij, M. G., Majorova, S. P., Vorobjev, I. A., Jandaghi, P., Bure, I. V., & Hoheisel, J. D. (2011). Correction of PCR-bias in quantitative DNA methylation studies by means of cubic polynomial regression. Nucleic Acids Research, 39(11). https://doi.org/10.1093/nar/gkr213

Correction of PCR-bias in quantitative DNA methylation studies by means of cubic polynomial regression. / Moskalev, Evgeny A.; Zavgorodnij, Mikhail G.; Majorova, Svetlana P.; Vorobjev, Ivan A.; Jandaghi, Pouria; Bure, Irina V.; Hoheisel, Jörg D.

In: Nucleic Acids Research, Vol. 39, No. 11, 06.2011.

Research output: Contribution to journalArticle

Moskalev, Evgeny A. ; Zavgorodnij, Mikhail G. ; Majorova, Svetlana P. ; Vorobjev, Ivan A. ; Jandaghi, Pouria ; Bure, Irina V. ; Hoheisel, Jörg D. / Correction of PCR-bias in quantitative DNA methylation studies by means of cubic polynomial regression. In: Nucleic Acids Research. 2011 ; Vol. 39, No. 11.
@article{f2b4ac4614324739865d138f8b5dcb03,
title = "Correction of PCR-bias in quantitative DNA methylation studies by means of cubic polynomial regression",
abstract = "DNA methylation profiling has become an important aspect of biomedical molecular analysis. Polymerase chain reaction (PCR) amplification of bisulphite-treated DNA is a processing step that is common to many currently used methods of quantitative methylation analysis. Preferential amplification of unmethylated alleles - known as PCR-bias - may significantly affect the accuracy of quantification. To date, no universal experimental approach has been reported to overcome the problem. This study presents an effective method of correcting biased methylation data. The procedure includes a calibration performed in parallel to the analysis of the samples under investigation. DNA samples with defined degrees of methylation are analysed. The observed deviation of the experimental results from the expected values is used for calculating a regression curve. The equation of the best-fitting curve is then used for correction of the data obtained from the samples of interest. The process can be applied irrespective of the locus interrogated and the number of sites analysed, avoiding an optimization of the amplification conditions for each individual locus.",
author = "Moskalev, {Evgeny A.} and Zavgorodnij, {Mikhail G.} and Majorova, {Svetlana P.} and Vorobjev, {Ivan A.} and Pouria Jandaghi and Bure, {Irina V.} and Hoheisel, {J{\"o}rg D.}",
year = "2011",
month = "6",
doi = "10.1093/nar/gkr213",
language = "English",
volume = "39",
journal = "Nucleic Acids Research",
issn = "0305-1048",
publisher = "Oxford University Press",
number = "11",

}

TY - JOUR

T1 - Correction of PCR-bias in quantitative DNA methylation studies by means of cubic polynomial regression

AU - Moskalev, Evgeny A.

AU - Zavgorodnij, Mikhail G.

AU - Majorova, Svetlana P.

AU - Vorobjev, Ivan A.

AU - Jandaghi, Pouria

AU - Bure, Irina V.

AU - Hoheisel, Jörg D.

PY - 2011/6

Y1 - 2011/6

N2 - DNA methylation profiling has become an important aspect of biomedical molecular analysis. Polymerase chain reaction (PCR) amplification of bisulphite-treated DNA is a processing step that is common to many currently used methods of quantitative methylation analysis. Preferential amplification of unmethylated alleles - known as PCR-bias - may significantly affect the accuracy of quantification. To date, no universal experimental approach has been reported to overcome the problem. This study presents an effective method of correcting biased methylation data. The procedure includes a calibration performed in parallel to the analysis of the samples under investigation. DNA samples with defined degrees of methylation are analysed. The observed deviation of the experimental results from the expected values is used for calculating a regression curve. The equation of the best-fitting curve is then used for correction of the data obtained from the samples of interest. The process can be applied irrespective of the locus interrogated and the number of sites analysed, avoiding an optimization of the amplification conditions for each individual locus.

AB - DNA methylation profiling has become an important aspect of biomedical molecular analysis. Polymerase chain reaction (PCR) amplification of bisulphite-treated DNA is a processing step that is common to many currently used methods of quantitative methylation analysis. Preferential amplification of unmethylated alleles - known as PCR-bias - may significantly affect the accuracy of quantification. To date, no universal experimental approach has been reported to overcome the problem. This study presents an effective method of correcting biased methylation data. The procedure includes a calibration performed in parallel to the analysis of the samples under investigation. DNA samples with defined degrees of methylation are analysed. The observed deviation of the experimental results from the expected values is used for calculating a regression curve. The equation of the best-fitting curve is then used for correction of the data obtained from the samples of interest. The process can be applied irrespective of the locus interrogated and the number of sites analysed, avoiding an optimization of the amplification conditions for each individual locus.

UR - http://www.scopus.com/inward/record.url?scp=79959393888&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=79959393888&partnerID=8YFLogxK

U2 - 10.1093/nar/gkr213

DO - 10.1093/nar/gkr213

M3 - Article

VL - 39

JO - Nucleic Acids Research

JF - Nucleic Acids Research

SN - 0305-1048

IS - 11

ER -