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- W275293624 abstract "AIM:To optimize conditions of DHPLC and analyze the effectiveness of various DNA polymerases on DHPLC resolution,and evaluate the sensitivity of DHPLC in the mutation screening of mitochondrial DNA(mtDNA).METHODS:Two fragments of 16s gene of mitochondrial DNA(one of them F2 is a mutant fragment) and an A3243G mutated fragment were used to analyze the UV detection limit and determine the minimum percentage of mutant PCR products for HDPLC and evaluate effcets of DNA polymerases on resolution of DHPLC.Under the optimal conditions.we analyzed the mtDNA mutations from muscle tissues of mitochondrial encephalomyopathy with lactic acidosis and stroke-like episodes(MELAS) and screened blindly for variances in D-loop region of mtDNA from human gastric tumor specimen.RESULTS:Teh A3243G variants were detected in 12 cases of MELAS,no alterations were detected in controls and these results were consistent with the results obtained by analysis of RFLP with APal.We also identified 26 D-loop variances in 46 cases of human gastric cancer tissues and 38 alterations in 13 gastric cancer cell lines.The mutation of mtDNA at 80ng PCR products containing a minimum of 5% mutant sequences could be detected by using DHPLC with UV detector Moreover,Ampli-Taq Gold polymerase was equally as good as the proofreading DNA polymerase(e.g.Pfu)in eliminating the false positive produced by Tay DNA polymerases.CONCLUSION:DHPLC is a powerful,rapid and sensitive mutaion screening method for mtDNA.Proofreading DNA Ploymerase in more suitalbe for HDPLC analysis than Taq polymerase." @default.
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- W275293624 date "2002-01-01" @default.
- W275293624 modified "2023-09-28" @default.
- W275293624 title "Rapd screening mitochondrial DNA mutation by using denaturing high—performance liquid chromatography" @default.
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