Identification of Base Pairs in Single-Nucleotide Polymorphisms by MutS Protein-Mediated Capillary Electrophoresis

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• 作者：Andrei P. Drabovich and Sergey N. Krylov
• 刊名：Analytical Chemistry
• 出版年：2006
• 出版时间：March 15, 2006
• 年：2006
• 卷：78
• 期：6
• 页码：2035 - 2038
• 全文大小：126K
• 年卷期：v.78,no.6(March 15, 2006)
• ISSN：1520-6882

文摘

Single-nucleotide polymorphisms (SNPs) are widespreadgenomic variations, which are associated with serioushealth disorders and drug resistance. Multiple clinicalapplications and studies of global population geneticsrequire fast and informative analysis of SNPs. Most ofconventional methods sense the presence of the SNP butcannot identify the base pair in it. Here we report simpleidentification of base pairs in SNPs without DNA sequencing. Our approach is based on the unique ability of MutSprotein to bind different single-nucleotide mismatches inDNA with different affinities. Conceptually, the DNA inquestion is mixed with reference DNA, melted, andreannealed. If the DNA in question has an SNP, theproducts of reannealing will have two different single-nucleotide mismatches, which provide a base-pair-specificsignature of the SNP. The products of reannealing aremixed with MutS, equilibrated, and separated by equilibrium capillary electrophoresis of equilibrium mixtureswith MutS in the run buffer. The pattern of migration timesof DNAs with mismatches is used for unequivocal identification of the base pair in the SNP. In addition to itsability to identify base pairs in SNPs, the new analyticalapproach is fast, simple, highly sensitive, and requiresno quantitation. It will find applications in studies ofheterogeneity of base pairs in known SNPs in large humanpopulations.