摘要
为构建胸苷激酶(Thymidine kinase,TK)基因缺失活载体疫苗,对山羊痘病毒(Goatpox virus,GPV)疫苗株(AV41)ORF64～ORF67进行了克隆和序列分析。结果表明:在全长3460bp的DNA序列中,包含4个完整的开放阅读框(Open reading frame,ORF)。ORF64核苷酸序列全长396bp,编码病毒膜蛋白;ORF65核苷酸序列全长444bp,ORF64和ORF65有44个碱基重叠。ORF66核苷酸序列全长534bp,编码胸苷激酶,具有保守的ATP结合位点和细胞中TK特征序列。ORF67核苷酸序列全长594bp,编码宿主范围相关蛋白。ORF64～ORF66在脊索动物痘病毒亚科中是完全保守的。与参考的国外GPV进行序列比较,ORF64～ORF67有一些差异,如突变和插入等。同源性分析显示:与羊痘病毒属比较,核苷酸和氨基酸同源性均较高(94.7%～100%)。我国的GPV不同毒株TK同源性为100%。与脊索动物痘病毒亚科中其他成员同源性差异较大(17.3%～65.2%)。将GPV AV41 TK与鸡、小鼠和人类的TK氨基酸序列进行比较,分析GPV AV41 TK进化关系表明,GPVTK基因在进化上可能起源于宿主细胞的TK1基因。本研究结果显示,在分子水平上我国GPV AV41与国外毒株存在差异,推测可能是GPV AV41疫苗株在致弱过程中发生一定程度的变异。
A goatpox virus （GPV） vaccine vector that has thymidine kinase （TK） deleted was constructed. DNA fragment spanning ORF64-ORF67 of GPV vaccine strain AV41 were amplified by polymerase chain reaction （PCR）, cloned into plasmid vector and sequenced. The result revealed a DNA fragment of 3 460 bp that contains four complete open reading frames （ORFs）. ORF64 of 396 bp encodes a membrane protein. ORF65 of 444 bp with 44 bp overlapping with ORF64 also encodes amembrane protein. ORF66 of 534 bp encodes TK protein and harbours a conserved ATP-binding motif （P-loop） and thymidine kinase cellular-type signature. ORF67 of 594 bp encodes a host range protein. ORF64-ORF66 were completely conserved in Chordopoxrinae （ChPV）. Compared to foreign GPV, namely GPV G20-LKV and Pellor strain, the ORF64-ORF67 regions contain some mutations and insertions. Sequence comparison analysis of 40RFs showed that GPV, sheeppox virus （SPV） and Lumpy skin disease virus （LSDV） shared highly conserved nucleotide and amino acid sequences, with 94.7 %-100 % homology. The amino acid sequences among GPV strains in China are identical, but differ from other poxvirus in Chordopoxrinae, with only 17.3 % to 65.2 % homology. In addition, analysis of phylogenetic relationship of GPV AV41 TK suggested that GPV TK appeared to derive from TK1 of the infected-cell.
出处
《中国预防兽医学报》
CAS
CSCD
北大核心
2007年第2期96-102,111,共8页
Chinese Journal of Preventive Veterinary Medicine
关键词
山羊痘病毒
胸苷激酶
分子特征
goat poxvirus
thymidine kinase （TK）
molecular characterization
