猪博卡病毒不同基因型三重PCR检测方法的建立及初步应用  

Development of a Triple PCR Method for Identifying Three Genotypes of Porcine bocavirus

作　　者：宏春慧 杨兵[2] 覃湘婕[2] 周宏专[2] 徐福洲[2] 苏霞[2] 薛静[3] 张晓东[3] 王金洛[2] 

机构地区：[1]北京农学院动物科学技术学院,北京102206 [2]北京市农林科学院畜牧兽医研究所畜禽疫病防控技术北京市重点实验室,北京100097 [3]北京农业生物技术研究中心,北京100097

出　　处：《华北农学报》2016年第2期12-16,共5页Acta Agriculturae Boreali-Sinica

基　　金：北京市农林科学院科技创新能力建设专项(KJCX20140409);北京市科技计划项目(Z131100003113015)

摘　　要：为鉴别猪博卡病毒1型、2型和3型3种基因型,根据猪博卡病毒基因序列分别设计3对针对保守区域的引物进行PCR扩增,扩增目的片段大小分别为645 bp(PBoV1型)、352 bp(PBoV2型)和259 bp(PBoV3型)。通过引物浓度和退火温度等条件优化,首次建立了同时检测猪博卡病毒3种基因型的三重PCR方法。所建立的三重PCR方法扩增其他猪病病毒结果均为阴性,最低检测限为8×10-7ng/μL。结果表明,该方法具有较好的特异性和敏感性。对临床样品进行三重PCR和单项PCR检测,对比结果显示符合率为100%。该方法的建立为猪博卡病毒3种不同基因型的鉴别检测提供了有效手段。To develop a method for identifying three genotypes of Porcine bocavirus,3 pairs of primers against conserved regions were designed according to the published gene sequences of Porcine bocavirus,and the PCR products were 645 bp（ PBoV1）,352 bp（ PBoV2）,259 bp（ PBoV3）,respectively. By optimizing the reaction conditions,a triple PCR method for detecting all three genotypes of Porcine bocavirus was established for the first time. Results showed that no specific band was amplified from other porcine viruses by the triple PCR. The detection limit of the method was 8 × 10- 7ng / μL. The results indicated that this method had not only good sensitivity but also high specificity. The clinical samples were used to detect three genotypes of Porcine bocavirus by the triple PCR and single PCR respectively with the coincidence of 100%. The establishment of the triple PCR method is helpful for identifying the three genotypes of Porcine bocavirus.

关 键 词：猪博卡病毒 基因型 三重PCR 检测 

分 类 号：Q78[生物学—分子生物学] S436[农业科学—农业昆虫与害虫防治]