文章摘要
Mei Hong,Enben Su,Ziqing Chen,Xiaobing Ju,Qi Chen,Rong Zhou.[J].南京医科大学学报,2008,28(6):346~350
Discrimination of mitochondrial DNA 10400 locus by SNP-operated on/off Switch
投稿时间:2008-10-10  
DOI:10.7655
中文关键词: 
英文关键词: SNP  exo+ polymerase  phosphorothioate-modification  allele-specific PCR  mitochondrial DNA
基金项目:
作者单位
Mei Hong The expert evidence institute, the First Affiliated Hospital of Nanjing Medical University, Nanjing 210029,Jiangsu Province, China 
Enben Su  
Ziqing Chen  
Xiaobing Ju  
Qi Chen  
Rong Zhou  
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中文摘要:
      
英文摘要:
      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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