猪瘟病毒国内分离株(HL-LY)E2基因的克隆及序列分析

用猪肾细胞（pK-15)繁殖猪瘟病毒（CSFV）分离株HL－LY，根据基因库已发表的CSFV E2基因序列，设计并合成一对引物，以CSFV总RNA为模板，通过RT－PCR技术扩增出约1.1kb的片段，即E2基因。将RT－PCR产物克隆到pMD18-T载体上，构建了重组质粒T－E2，经酶切、PCR鉴定和序列测定，表明均已成功获得了CSFV E2基因的重组体T－E2。将该测序结果与国内几个毒株SHIMEN，C，C－V－LZ,GS-LT,GS-LX德育了列同源性比较，核苷酸同源性分别为96.59%,96.005,88.495,78.24%,77.82%；氨基酸同源性分别为99.46%,98.39%,93.54%,90.70%,90.44%。表明该毒株与国内标准毒株SHIMEN株和C株具有较高的同源性。

Cloning and Sequence Analysis of E2 Gene from Domestic Isolate Strain of CSFV

The HL-LY strain of Classical Swine Fever Virus(CSFV) was propagated and harvested on pk-15.One pair of primers were designed to amplify E 2 gene by RT-PCR according to published sequence of CSFV's E 2 gene of gene bank . The product of RT-PCR,which is about 1.1 kb was cloned into pMD18-T vector,then the recombinent plasmid which is named T-E 2 was identified by corresponding restriction endonuclease and PCR and was analyzed with sequencing and homology compariation.The result showed that CSFV E 2 gene of HL-LY strain share 96.59%,96.00%,88.49%,78.24%,77.82% with SHIMEN strain,C strain,C-V-LZ strain , GS-LT strain,GS-LX strain in nucleotide sequence respectively, and share 99.46%,98.39%,93.54%,90.70%,90.44% identity in amino acids sequence respectively,which showed that HL-LY strain has high homology with domestic standard SHIMEN strain and C strain.