Restriction site-specific methylation studies of imprinted genes with quantitative real-time PCR.

Clinical Chemistry
Sara BruceJuha Kere

Abstract

Epigenetic studies, such as the measurement of DNA methylation, are important in the investigation of syndromes influenced by imprinted genes. Quick and accurate quantification of methylation at such genes can be of appreciable diagnostic aid. We first digested genomic DNA with methylation-sensitive restriction enzymes and used DNA without digestion as a control and nonmethylated lambda DNA as an internal control for digestion efficiency. We then performed quantitative real-time PCR analyses with 6 unique PCR assays to investigate 4 imprinting control regions on chromosomes 7 and 11 in individuals with uniparental disomy of chromosome 7 (UPD7) and in control individuals. Our validation of the method demonstrated both quantitative recovery and low methodologic imprecision. The imprinted loci on chromosome 7 behaved as expected in maternal UPD7 (100% methylation) and paternal UPD7 (<10% methylation). In controls, the mean (SD) for percent methylation at 2 previously well-studied restriction sites were 46% (6%) for both H19 and KCNQ1OT1, a result consistent with the previously observed methylation rate of approximately 50%. The methylation percentages of all investigated imprinted loci were normally distributed, implying that the ...Continue Reading

References

Aug 1, 1994·Human Molecular Genetics·W ReikE R Maher
Jun 15, 1997·Nucleic Acids Research·Z Xiong, P W Laird
Jul 8, 1999·Proceedings of the National Academy of Sciences of the United States of America·N J SmilinichM J Higgins
Oct 3, 1999·The Journal of Biological Chemistry·M A FrevelA E Reeve
May 23, 2001·Journal of Medical Genetics·J F Costello, C Plass
Feb 16, 2002·Methods : a Companion to Methods in Enzymology·B N TrinhP W Laird
Sep 20, 2002·BioTechniques·Mario F Fraga, Manel Esteller
Mar 15, 2003·Science·Hengmi CuiAndrew P Feinberg
Jan 15, 2004·Nucleic Acids Research·Susan E CottrellKurt Berlin
Dec 4, 2004·Nucleic Acids Research·Hélène ThomassinThierry Grange
May 10, 2005·Journal of Medical Genetics·S BruceJ Kere
Jul 2, 2005·Trends in Genetics : TIG·Ian M MorisonHamish G Spencer
Jul 7, 2005·European Journal of Human Genetics : EJHG·Wendy N CooperEamonn R Maher
Oct 26, 2005·Proceedings of the National Academy of Sciences of the United States of America·Mathias EhrichDirk van den Boom
Apr 1, 2006·Cytogenetic and Genome Research·B Horsthemke, K Buiting
Apr 7, 2007·Pediatric Research·Adam C SmithRosanna Weksberg

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Citations

Dec 29, 2012·Genetics and Molecular Biology·Leila C A CardosoFernando R Vargas
Nov 9, 2010·Taiwanese Journal of Obstetrics & Gynecology·Chih-Ping ChenWayseen Wang
Apr 5, 2011·Fertility and Sterility·Maria SandbackaHannele Laivuori
Oct 13, 2009·Experimental and Molecular Pathology·Jean-François GagnonCaroline Diorio
May 18, 2010·Clinical Chemistry·Thomas von KanelSabina Gallati
Sep 2, 2016·Clinical Cancer Research : an Official Journal of the American Association for Cancer Research·Hye-Young MinHo-Young Lee

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