High resolution melting curve analysis as a new tool for rapid identification of canine parvovirus type 2 strains

文献类型： 外文期刊

第一作者： Bingga, Gali

作者： Bingga, Gali;Ding, Shuangyang;Bingga, Gali;Liu, Zhicheng;Zhang, Jianfeng;Lin, Lifeng;Zhu, Yujun;Guo, Pengju

作者机构：

关键词： PCR;Canine parvovirus;VP2 gene;High resolution melting curve analysis;Heteroduplex

期刊名称：MOLECULAR AND CELLULAR PROBES （ 影响因子：2.365； 五年影响因子：2.386 ）

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收录情况： SCI

摘要： A high resolution melting (HRM) curve method was developed to identify canine parvovirus type 2 (CPV-2) strains by nested PCR. Two sets of primers, CPV-426F/426R and CPV-87R/87F, were designed that amplified a 52 bp and 53 bp product from the viral VP2 capsid gene. The region amplified by CPV-426F/426R included the A4062G and T4064A mutations in CPV-2a, CPV-2b and CPV-2c. The region amplified by CPV-87F/87R included the A3045T mutation in the vaccine strains of CPV-2 and CPV-2a, CPV-2b and CPV-2c. Faecal samples were obtained from 30 dogs that were CPV antigen-positive. The DNA was isolated from the faecal samples and PCR-amplified using the two sets of primers, and genotyped by HRM curve analysis. The PCR-HRM assay was able to distinguish single nucleotide polymorphisms between CPV-2a, CPV-2b and CPV-2c using CPV-426F/426R. CPV-2a was distinguished from CPV-2b and CPV-2c by differences in the melting temperature. CPV-2b and CPV-2c could be distinguished based on the shape of the melting curve after generating heteroduplexes using a CPV-2b reference sample. The vaccine strains of CPV-2 were identified using CPV-87F/87R. Conventional methods for genotyping CPV strains are labor intensive, expensive or time consuming; the present PCR-based HRM assay might be an attractive alternative. (C) 2014 Elsevier Ltd. All rights reserved.

分类号： R44

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