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Discrimination of mitochondrial DNA 10400 locus by SNP-operated on/off Switch
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This work was supported by a grant from the"135"Foundation of Jiangsu Province(RC2002052)

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    Objective:To apply reformed AS-PCR, which combined phosphorothioate-modified primers with exo+ polymerase, in single nucle-otide polymorphism discrimination of mitochondrial DNA 10400 locus. Methods: We used the mtDNA 10400 locus to design unmodi-fied and 3′phosphorothioate-modified allele-specific primers for PCR, which was performed using polymerases with and without 3′exonuclease activities. The effects of these primers on primer-extension were evaluated by agarose gel electrophoresis. Results: The unmodified primers were extended by both exo- and exo+ polymerase irrespective of whether the primers were matched or mismatched with the templates. However, the 3′phosphorothioate-modified primers with a terminal mismatch triggered an “off-switch”of exo+ polymerase when compared to exo-polymerase. Conclusion: The“on/off”switch constituted by the combination of 3′phosphorothioate-modified primers with exo+ polymerase is a cost-effective, high-throughput and reliable method for SNP typing, which will be of enormous application in association studies by single nucleotide polymorphism screening.

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Mei Hong, Enben Su, Ziqing Chen, Xiaobing Ju, Qi Chen, Rong Zhou.[J].南京医科大学学报(自然科学版),2008,28(6):346-350

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  • 收稿日期:2008-10-10
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