PMID: 16526411Mar 11, 2006Paper

Two-color quantitative multiplex methylation-specific PCR

BioTechniques
Theresa Swift-ScanlanMary Jo Fackler

Abstract

In recent years, several methylation-specific PCR-based techniques have been developed to identify and characterize hypermethylation of CpG dinucleotides with the primary goal of elucidating a better understanding of the role of DNA methylation in important biological processes, such as chromosome X inactivation and carcinogenesis. The specificity of methylation-specific PCR (MSP) techniques relies on amplifying sodium bisulfite-treated DNA with primers specific to predicted sequences of unmethylated and methylated DNA within the gene of interest. In the past, unmethylated and methylated reactions were singleplex and performed in separate wells. In this paper we report a modification of the real-time quantitative multiplex MSP (QM-MSP) technique of Fackler and colleagues that can be applied to any real-time MSP experiment. Although co-amplification with multiple fluorophores is common in standard reverse transcription PCR (RT-PCR), MSP presents unique challenges both mechanistically and operationally that must be overcome in order to successfully co-amplify two methylation-specific targets. In this two-color modification, unmethylated and methylated primer/probe sets are successfully co-amplified in the same reaction using FAM-...Continue Reading

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Citations

Dec 17, 2008·Pharmacogenomics·Pang-Kuo Lo, Saraswati Sukumar
Aug 13, 2010·Biomarkers in Medicine·Leonel Maldonado, Mohammad Obaidul Hoque
Nov 9, 2011·Chinese Journal of Cancer·Xiao-Tong Hu, Chao He
Aug 28, 2009·The Journal of Molecular Diagnostics : JMD·Pang-Kuo LoSaraswati Sukumar
May 25, 2010·Virchows Archiv : an International Journal of Pathology·Jo-Heon KimSaraswati Sukumar
Oct 16, 2015·Tumour Biology : the Journal of the International Society for Oncodevelopmental Biology and Medicine·Sandra MersakovaErika Halasova
Jan 7, 2010·Cancer Prevention Research·Wendy Wang, Sudhir Srivastava
May 31, 2013·Clinical Cancer Research : an Official Journal of the American Association for Cancer Research·Vered StearnsNancy E Davidson

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