pcr熔解曲线法检测hbv基因型及rtarmsqpcr法定量检测hbvymdd突变建立-detection of hbv genotype by pcr melting curve method and establishment of hbv ymdd mutation by rtarmsqpcr method.docxVIP

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pcr熔解曲线法检测hbv基因型及rtarmsqpcr法定量检测hbvymdd突变建立-detection of hbv genotype by pcr melting curve method and establishment of hbv ymdd mutation by rtarmsqpcr method.docx

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pcr熔解曲线法检测hbv基因型及rtarmsqpcr法定量检测hbvymdd突变建立-detection of hbv genotype by pcr melting curve method and establishment of hbv ymdd mutation by rtarmsqpcr method

Genotyping of Hepatitis B Virus by Melting Curve Analysis and Quantitative Detection of Hepatitis B Virus YMDD Motif Mutation by Amplification Refractory MutationSystem Combined with SYBR GreenⅠAbstractObjective:To develop a new method for hepatitis B virus(HBV) genotyping, and detect the HBV genotypes of some patients who are infected with HBV, so that we can investigate the distribution of HBV genotypes in Fujian Province, and discuss the relationship between HBV genotypes with liver fibrosis, HBeAg positive incidence, and HBV DNA level.Construct plasmids of YMDD, YVDD, to establish and evaluate the method for detecting HBV YMDD motif mutation quantitatively by amplification refractory mutation system (ARMS).Methods:The establishment of method for HBV genotyping by PCR melt curve analysis: We designed type-specific primers according to the analysis of HBV B and C genomes in China reported in document and use HBV B and C plasmids as standard, to establish a new method for detecting HBV genotype with melt curve analysis based on real-time PCR, we could get the HBV genotype by Tm value of PCR product. Evaluate its sensitivity, specificity and repetitiveness.The evaluation of method for HBV genotyping by PCR melt curve analysis: Detect HBV genotypes of 51 patients who are infected with HBV by melt curve analysis and kit used for HBV genotyping on market respectively, and compare the coincidence rate of two assays to evaluate the melt curve analysis.The clnical application of method for HBV genotyping by PCR melt curve analysis: Detect HBV genotypes of 346 patients who is HBsAg-positive with the meltcurve analysis to investigate the distribution of HBV genotypes in Fujian Province, and discuss the relationship between the HBV genotype with liver fibrosis, HBeAg positive incidence, and HBV DNA level.The establishment of method for detecting HBV YMDD motif mutation by RT-ARMS-qPCR: YMDD and YVDD plasmids were constructed and used as standard to establish a stand cu