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filingDate 2005-10-11-04:00^^<http://www.w3.org/2001/XMLSchema#date>
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publicationDate 2006-05-24-04:00^^<http://www.w3.org/2001/XMLSchema#date>
publicationNumber EP-1659186-A1
titleOfInvention A method for the carry-over protection in DNA amplification systems targeting methylation analysis achieved by a modified pre-treatment of nucleic acids
abstract Disclosed is a method for the specific amplification of template DNA in the presence of potentially contaminating PCR products from previous amplification experiments. In the first step DNA is contacted with a bisulfite solution, which reacts with unmethylated cytosines but not with methylated cytosines, by sulfonating them. This results in deamination of the cytosine whereby sulfonated uracil is generated. Such sulfonation protects the template nucleic acid from being a target for the enzyme UNG. Any contaminating DNA, which contains unprotected unsulfonated or desulfonated uracils is degraded enzymatically while UNG is active. After UNG treatment and inactivation, the sulfonated uracil bases are converted into uracil by desulfonation. This method is useful for decontamination of nucleic acid samples, or rather for avoiding amplification of 'carry over products' in particular in the context of DNA methylation analysis. Furthermore it can be used as a simplified method of bisulfite treatment in gerneral.
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