应用巢式PCR检测鸭圆环病毒浙江株及其全基因序列分析

Nested PCR detection and sequence analysis on Zhejiang duck circovirus

应用巢式PCR(nested PCR)技术对收集自浙江温州种番鸭进行鸭圆环病毒核酸扩增,并对测定的序列进行分析,结果用套式引物扩增出目的条带片段大小约为340 nt。根据序列测定结果设计1对反向引物用于测定病毒基因组余下的基因片段。序列分析表明该圆环病毒基因组全长为1995 nt,其ORF-V1编码292个氨基酸,ORF-C1编码257个氨基酸。序列分析表明,本试验株与番鸭分离株(GenBank登录号EF451157)和半番鸭分离株(GenBank登录号EU344804)的同源性最高,均高达99.2%,与鸭圆环病毒欧洲(德国)分离株和北美(美国)分离株在同一支遗传进化关系上。

Using the nested PCR technology, the duck circovirus （DuCV） nucleotide fragment, which was isolated from Wenzhou, Zhejiang, was amplified and its sequence analyzed. The result showed that the obtained segment was about 340nt in length. Based on the determined sequence, two primers were applied for the construction of the remaining genomic sequence by inverse PCR. The genome of the Zhejiang DuCV had a length of 1995nt. Its ORF-V1 encoded a replication protein of 292 amino acids, while the ORF-C1 encoded a capsid protein of 257 amino acids. The strain had highest sequence similarities （i. e. , 99.2%） with EF451157 detected from the muscovy duck and EU344804 detected from the mule duck in Fujian. Phylogenetic analysis with Lasergene v 7. 1 DNAStar software by Clustal W method showed that it had the same sub-lineage with the European strain detected in German mulard duck and the North American strain found in the American Perkin duck.