单纯疱疹病毒Ⅰ型胸苷激酶基因的扩增、克隆和测序

目的：对单纯疱疹Ⅰ型胸苷激酶基因扩增、克隆和测序．方法：从单纯疱疹病毒Ⅰ型（ＨＳＶ－１）１７ｓｙｎ＋株感染的ＢＨＫ２１细胞上清液中提取ＨＳＶ－１基因组ＤＮＡ，以此为模板，用ＰＣＲ方法扩增胸苷激酶（ＴＫ）．将扩增的片段克隆入载体ｐＵＣ１８中，并进行测序．结果：除终止码外，ＨＳＶ－１ＴＫ基因全部编码区为１１２８ｂｐ，编码３７６个氨基酸，其中含１２个蛋氨酸，４个半胱氨酸．ＴＫ的第２４９和２５０位氨基酸残基分别为Ｌｅｕ和Ｇｌｎ，其相应密码子为ＣＴＧ，ＧＡＧ，构成１个ＰｓｔＩ位点，第３１３和３１４氨基酸残基分别为Ａｓｐ和Ｖａｌ，其相应密码子为ＧＡＣ和ＧＴＣ，构成另一个ＰｓｔＩ位点．结论：本研究扩增出了ＨＳＶ－１ＴＫ基因的全部编码区序列

Amplification,cloning and sequencing of the thymidine kinase gene of herpes simplex virus I

Aim: To amplificate,clone and sequence the thymidine kinase (TK) gene of herpes simplex virus 1(HSV I). Methods:The BHK21 cells were infected by HSV I 17 syn + strain. HSV I genomic DNA was purified from the BHK 21 cells suspension and used as template to run PCR for TK gene amplification. The amplificated products were cloned into pUC 18 vector and sequenced. Results: Coding region of HSV 1 TK gene consisted of 1128 bp except stop code, it encoded 376 amino acids. Among the 1128 bp, there were 12 Met and 4 Cys. The 249 and 250 amino acid residues of TK were Leu and Gln respectively. Their corresponding codes were CTG and GAG which formed one Pst1 site. The 313 and 314 amino acid residues were Asp and Val respectively. Their corresponding codes were GAC and GTC which formed another PstI site. Conclusion:The coding sequence amplificated in this study is the entire sequence of coding region of HSV 1 TK.