Combination of methylated-DNA precipitation and methylation-sensitive restriction enzymes (COMPARE-MS) for the rapid, sensitive and quantitative detection of DNA methylation
Identifieur interne : 001673 ( Main/Exploration ); précédent : 001672; suivant : 001674Combination of methylated-DNA precipitation and methylation-sensitive restriction enzymes (COMPARE-MS) for the rapid, sensitive and quantitative detection of DNA methylation
Auteurs : Srinivasan Yegnasubramanian [États-Unis] ; Xiaohui Lin [États-Unis] ; Michael C. Haffner [États-Unis, Autriche] ; Angelo M. Demarzo [États-Unis] ; William G. Nelson [États-Unis]Source :
- Nucleic Acids Research [ 0305-1048 ] ; 2006.
Abstract
Hypermethylation of CpG island (CGI) sequences is a nearly universal somatic genome alteration in cancer. Rapid and sensitive detection of DNA hypermethylation would aid in cancer diagnosis and risk stratification. We present a novel technique, called COMPARE-MS, that can rapidly and quantitatively detect CGI hypermethylation with high sensitivity and specificity in hundreds of samples simultaneously. To quantitate CGI hypermethylation, COMPARE-MS uses real-time PCR of DNA that was first digested by methylation-sensitive restriction enzymes and then precipitated by methyl-binding domain polypeptides immobilized on a magnetic solid matrix. We show that COMPARE-MS could detect five genome equivalents of methylated CGIs in a 1000- to 10 000-fold excess of unmethylated DNA. COMPARE-MS was used to rapidly quantitate hypermethylation at multiple CGIs in >155 prostate tissues, including benign and malignant prostate specimens, and prostate cell lines. This analysis showed that GSTP1, MDR1 and PTGS2 CGI hypermethylation as determined by COMPARE-MS could differentiate between malignant and benign prostate with sensitivities >95% and specificities approaching 100%. This novel technology could significantly improve our ability to detect CGI hypermethylation.
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DOI: 10.1093/nar/gnj022
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Rapid and sensitive detection of DNA hypermethylation would aid in cancer diagnosis and risk stratification. We present a novel technique, called COMPARE-MS, that can rapidly and quantitatively detect CGI hypermethylation with high sensitivity and specificity in hundreds of samples simultaneously. To quantitate CGI hypermethylation, COMPARE-MS uses real-time PCR of DNA that was first digested by methylation-sensitive restriction enzymes and then precipitated by methyl-binding domain polypeptides immobilized on a magnetic solid matrix. We show that COMPARE-MS could detect five genome equivalents of methylated CGIs in a 1000- to 10 000-fold excess of unmethylated DNA. COMPARE-MS was used to rapidly quantitate hypermethylation at multiple CGIs in >155 prostate tissues, including benign and malignant prostate specimens, and prostate cell lines. This analysis showed that GSTP1, MDR1 and PTGS2 CGI hypermethylation as determined by COMPARE-MS could differentiate between malignant and benign prostate with sensitivities >95% and specificities approaching 100%. This novel technology could significantly improve our ability to detect CGI hypermethylation.</div></front></TEI><affiliations><list><country><li>Autriche</li><li>États-Unis</li></country><region><li>Maryland</li></region></list><tree><country name="États-Unis"><region name="Maryland"><name sortKey="Yegnasubramanian, Srinivasan" sort="Yegnasubramanian, Srinivasan" uniqKey="Yegnasubramanian S" first="Srinivasan" last="Yegnasubramanian">Srinivasan Yegnasubramanian</name></region><name sortKey="Demarzo, Angelo M" sort="Demarzo, Angelo M" uniqKey="Demarzo A" first="Angelo M." last="Demarzo">Angelo M. Demarzo</name><name sortKey="Haffner, Michael C" sort="Haffner, Michael C" uniqKey="Haffner M" first="Michael C." last="Haffner">Michael C. Haffner</name><name sortKey="Lin, Xiaohui" sort="Lin, Xiaohui" uniqKey="Lin X" first="Xiaohui" last="Lin">Xiaohui Lin</name><name sortKey="Nelson, William G" sort="Nelson, William G" uniqKey="Nelson W" first="William G." last="Nelson">William G. Nelson</name><name sortKey="Nelson, William G" sort="Nelson, William G" uniqKey="Nelson W" first="William G." last="Nelson">William G. Nelson</name></country><country name="Autriche"><noRegion><name sortKey="Haffner, Michael C" sort="Haffner, Michael C" uniqKey="Haffner M" first="Michael C." last="Haffner">Michael C. Haffner</name></noRegion></country></tree></affiliations></record>Pour manipuler ce document sous Unix (Dilib)
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