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为建立可鉴定PCV2b和PCV2d基因亚型的PCR方法,通过分析Gen Bank内PCV2 ORF2基因序列,设计了可分别扩增ORF2基因和鉴别PCV2b、PCV2d的引物,并建立PCR方法。应用PCV2b和PCV2d引物扩增108份PCV2阳性样本,选择11份PCV2b和19份PCV2d单一阳性样本扩增ORF2基因序列,并用于遗传进化分析。结果显示,PCV2b和PCV2d引物扩增的敏感性较好,PCV2b和PCV2d最低可扩增基因拷贝数分别为19 copies/L和80 copies/L。108份PCV2阳性DNA中PCV2b阳性率为42.5%,PCV2d阳性率为69.4%,混合感染阳性率为12%。30条ORF2序列的遗传进化分析结果显示,11条为PCV2b基因亚型,19条为PCV2d基因亚型,遗传进化分析结果与PCR结果一致。本试验建立了一种鉴别PCV2b和PCV2d的PCR方法,可用于PCV2流行病学的调查。
To establish a PCR method that can identify PCV2b and PCV2d gene subtypes, primers for amplifying ORF2 gene and PCV2b and PCV2d were designed by analyzing the sequence of PCV2 ORF2 gene in GenBank, and PCR method was established. PCV2b and PCV2d primers were used to amplify 108 samples of PCV2 positive samples, select 11 copies of PCV2b and 19 copies of PCV2d single positive samples amplified ORF2 gene sequence and used for genetic analysis. The results showed that the primers of PCV2b and PCV2d amplified more sensitively, and the copies of PCV2b and PCV2d were 19 copies / L and 80 copies / L, respectively. The positive rate of PCV2b in PCV2 positive DNA was 42.5%, the positive rate of PCV2d was 69.4%, and the positive rate of mixed infection was 12%. Genetic evolution analysis of 30 ORF2 sequences showed that 11 were PCV2b genotypes and 19 were PCV2d genotypes. The results of genetic analysis were consistent with PCR results. This experiment established a PCR method to identify PCV2b and PCV2d, which can be used for epidemiological investigation of PCV2.