Constructin and Appraisement of Fusion Gene Eukaryon Expression VectorpcDNA3/HSVⅡ TK/Angiostatin_CHINESE JOURNAL OF BASES AND CLINICS IN GENERAL SURGERY

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Department of Hepatobiliary Surgery， The Second Affiliated Hospital of Kunming Medical College, Kunming 650101;

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Keywords：

Herpes simplex virus-Ⅱ; Thymidine kinase gene; Angiostation; Clone; Eukaryon expression vector; Fusion gene

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Objective To amplificate,clone and sequence the thymidine kinase (TK) gene of herpes simplex virusⅡ(HSVⅡ); to construct and appraise the fusion gene eukaryon expression vector, pcDNA3/HSVⅡ TK/angiostatin. MethodsThe Hep2 cells were infected by HSVⅡ Sav strain. HSVⅡ genomic DNA was purified from the Hep2 cells suspension and used as template to run PCR for TK gene amplification. The amplified products were cloned into PC DNA3 vector and sequenced. The vector pcDNA/HSVⅡ TK was cut by endonuclease. The gained TK gene was cloned into eukaryon expression vector. pcDNA3/angiostation, which had been constructed. ResultsCoding region of HSVⅡTK gene consisted of 1 128 bp except stop code, it encoded 376 amino acids.After cutting the new vector by endonuclease Hind Ⅲ and BamH Ⅰ,we gained the following gene fragment: 1000 bp (TK) and 700 bp (angiostation).Conclusion The fusion gene eukaryon expression vector， pcDNA3/HSVⅡ TK/angiostatin has been constructed.

Citation： WANG Lin,GUO Yongzhang,LI Li,et al.. Constructin and Appraisement of Fusion Gene Eukaryon Expression VectorpcDNA3/HSVⅡ TK/Angiostatin. CHINESE JOURNAL OF BASES AND CLINICS IN GENERAL SURGERY, 2002, 9(4): 246-248. doi: Copy

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1. Moolten FL.Tumor chemosensitivity conferred by inserted herpes thymidine kinase genes: paradigm for a prospective cancer control strategy [J]. Cancer Res,1986； 46(10)∶5276.
2. Nagy H, Panis Y, Fabre M, et al.Are hepatomas a good target for suicide gene therapy? An experimental study in rats using retroviralmediated transfer of thymidine kinase gene [J]. Surgery,1998; 123(1)∶ 19.
3. O’Reilly MS, Holmgren L, Shing Y,et al.Angiostatin: a novel angiogenesis inhibitor that mediates the suppression of metastasis by a Lewis lung carcinoma [J]. Cell,1994; 79(2)∶ 315.
4. Kit S, Kit M, Qavi H, et al. Nucleotide sequence of the herpes simplex virus type 2(HSV2)thymidine kinase gene and predicted amino acid sequence of thymidine kinase polypeptide and its comparison with the HSV1 thymidine kinase gene [J]. Biochim Biophys Acta,1983; 741(2)∶158.
5. 萨姆布鲁克J，弗里奇EF，曼尼阿蒂斯文T.分子克隆实验技术指南[M]. 第2版.北京：科学出版社，1992∶237~340.
6. Kuriyama S, Nakatani T, Masui K, et al. Bystander effect caused by suicide gene expression indicates the feasibility of gene therapy for hepatocellular carcinoma [J]. Hepatology,1995; 22(6)∶1838.
7. Gately S,Twardowski P,Stack MS,et al.The mechanism of cancermediated conversion of plasminogen to the angiogenesis inhibitor angiostatin [J]. Proc Natl Acad Sci USA,1997; 94(20)∶10868.
8. Sim BK,O’Reilly MS,Liang H,et al. A recombinant human angiostatin protein inhibits experimental primary and metastatic cancer [J]. Cancer Res, 1997; 57(7)∶1329.

CHINESE JOURNAL OF BASES AND CLINICS IN GENERAL SURGERY

Constructin and Appraisement of Fusion Gene Eukaryon Expression VectorpcDNA3/HSVⅡ TK/Angiostatin

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