伪狂犬病病毒HS株tk基因的PCR扩增与克隆

PCR Amplification and Cloning of tk Gene of Pseudorabies Virus HS 9304 Strain

参照伪狂犬病病毒（Ｐｓｅｕｄｏｒａｂｉｅｓｖｉｒｕｓ，ＰＲＶ）ＮＩＡ－３株ｔｋ基因序列，设计并合成１对长２２ｍｅｒ的引物，引物间距１．５ｋｂ，其内包含完整的ＰＲＶｔｋ基因。以ＢＨＫ２１细胞繁殖的ＰＲＶ湖北地方强毒株（ＨＳ－９３０４）基因组为模板进行ＰＣＲ扩增。琼脂糖凝胶电泳检测显示扩增主带清晰，长约１．５ｋｂ，符合设计要求。扩增片段克隆至由ｐＵＣ１８质粒改制而成的Ｔ载体中，限制性内切酶ＢａｍＨＩ、ＳｍａⅠ、ＸｈｏⅠ、ＨｉｎｄⅢ酶切分析证实，扩增片段的酶切位点与ｔｋ基因一致，说明扩增和克隆片段包含ＰＲＶｔｋ基因。

One pair of PCR primer for the PRV tk gene was designed according to the DNA sequence of PRV NIA 3 strain. With the primers, DNA fragment (about 1.5 kb) containing tk gene of PRV HS 9304 strain was successfully amplified by PCR. The amplified fragment was cloned into pUC18 plasmid. Restricted endonuclease analysis confirmed the accomplishment of the amplification and cloning of tk gene of PRV HS 9304 strain.