鸭瘟病毒强弱毒株TK基因的克隆及序列比较分析

根据GenBank中已发表的鸭瘟病毒TK基因序列,设计一对引物,对1株鸭瘟病毒强毒和1株鸭瘟病毒疫苗毒进行PCR扩增。将扩增的目的片段分别克隆到pMD18-T载体,经EcoRⅠ和HindⅢ双酶切鉴定,获得阳性重组质粒,然后对阳性重组质粒进行序列测定及分析。结果表明,本实验所扩增的鸭瘟病毒TK基因及侧翼UL24基因大小为1 995 bp,鸭瘟强、弱毒株TK基因及侧翼UL24基因序列完全相同,该病毒TK基因与鸭瘟病毒其它毒株AY911509与AY963569,四川株DQ640611,AV1221株EF173464,sd-01株EF417996同源性分别为99.5%,99.9%,100%,99.9%,100%,而该病毒UL24基因与鸭瘟病毒其它毒株AY911511,DQ227739,EF417996同源性为99.9%,99.8%,99.9%,表明鸭瘟病毒强弱毒株TK基因及侧翼UL24基因高度保守。为构建鸭瘟病毒TK基因缺失的转移载体奠定了基础。

Comparative Analysis on Clone and Sequence of TK Gene of Virulent and Vaccine DPV Strains

According to the previously described sequence of duck plague virus TK gene,one pair of primers was designed to amplify this gene in virulent and vaccine duck plague virus strains. The amplified products were purified and cloned into pMD18-T vector, and identified with the restriction enzyme EcoR I and Hind. The recombinant plasmids were sequenced and analysed,and a product of 1995 bp was obtained. This result indicated that the sequence of TK gene and UL24 was1995 bp. The sequence of virulent and vaccine duck plague virus strains was completely identical. Sequence analysis dates demonstrated that the DNA homology of virulent and vaccine duck plague virus strains shared 99. 5%,99.9%, 100%,99.9%, 100% similarities with the published sequences of AY911509 strains, AY963569 Strains, DQ640611 strains, AV1221 EF173464 Strains, sd 01 EF417996 Strains, while it shared 100%,99.8% ,99.9% similarities with the published sequences of UL24 AY911511 strains, DQ227739 strains, sd-01 EF417996 strains . The results indicated that TK gene and UL24 of duck plague virus viruses are highly genetically conserved. This study laid the foundation for the construction of transfer vector of deletion TK of duck plague virus.