新法制备含VEGF启动子驱动的CD/TK融合基因重组腺病毒

南方医科大学学报 ›› 2004, Vol. 24 ›› Issue (01): 117-120.

• • 上一篇

新法制备含VEGF启动子驱动的CD/TK融合基因重组腺病毒

黄元媛¹, 黄宗海¹, 陈建发¹, 汤福祥¹, 杨文宇¹, 郑全¹, 车小燕²

1. 第一军医大学珠江医院, 普通外科, 广东, 广州, 510282;
2. 第一军医大学珠江医院, 中心实验室, 广东, 广州, 510282

出版日期:2004-01-20 发布日期:2004-01-20
基金资助:
收稿日期:2003-6-28。
基金项目:国家“863”计划生物和现代农业技术研究基金(2001AA217171);广东省自然科学基金重点项目(013072)
作者简介:黄元媛(1978- ),女,2003年毕业于第一军医大学,硕士,医师,电话:020-61643211

An easier method for generating recombinant adenovirus containing CD/TK fusi on gene driv-en by vascular endothelial growth factor promoter

HUANG Yuan-yuan¹, HUANG Zong-hai¹, CHEN Jian-fa¹, TANG Fu-xiang¹, YANG Wen-yu¹, ZHENG Quan¹, CHE Xiao-yan²

1. 第一军医大学珠江医院, 普通外科, 广东, 广州, 510282;
2. 第一军医大学珠江医院, 中心实验室, 广东, 广州, 510282

Online:2004-01-20 Published:2004-01-20

摘要/Abstract

摘要： 目的应用简化的细菌内同源重组法高效制备含血管内皮细胞生长因子(VEGF)启动子驱动的CD/TK融合基因重组腺病毒。方法先以氯化钙法替代电穿孔法将腺病毒基因组质粒pAdEasy-1导入BJ5183细菌,制备BJ5183pAdEasy-1细菌;再以后者作感受态细菌,与线性化的转移质粒pAdtrack-VEGFP-CDglyTK进行同源重组,获得重组腺病毒质粒pAdEasy-VEGFP-CDglyTK,转染293细胞,制备含VEGF启动子驱动的CD/TK融合基因重组腺病毒。结果构建重组腺病毒质粒的成功率为60%(6/10)。转染293细胞后7~12d可收获病毒。结论简化的细菌内同源重组法简便易行,重组效率较高,易于推广应用。

Abstract: Objective To efficiently construct the recombinant adenov ir us containing CD/TK fusion gene driven by vascular endothelial growth factor (V EGF) promoter using improved homologous recombination in bacteria.Methods Chemi cal trans-formation, instead of electroporation, of the plasmid pAdEasy-1 into E.coli BJ5183 strain was performed to prepare BJ5183pAdEasy-1 as t he competent bacterium, in which pAdEasy-1 and pAdtrack-VEGFP-CDglyTK were recom bined. Fi-nally, the recombinant adenovirus of AdVEGFP-CDglyTK was constructed b y tranfecting 293 cells with linearized adenoviral genomes of pAdEasy-VEGFP-CDg lyTK.Results This new transformation procedure generated recombinant plasmids i n a yield of 60% (6/10), and the adenovirus AdVEGFP-CDglyTK was harvested 7-12 d after transfection.Conclusion The im-proved homologous recombination in bacte ria is efficient, convenient and easy to be carried out.

中图分类号:

Q784
R373.1

黄元媛¹, 黄宗海¹, 陈建发¹, 汤福祥¹, 杨文宇¹, 郑全¹, 车小燕². 新法制备含VEGF启动子驱动的CD/TK融合基因重组腺病毒[J]. 南方医科大学学报, 2004, 24(01): 117-120.

HUANG Yuan-yuan¹, HUANG Zong-hai¹, CHEN Jian-fa¹, TANG Fu-xiang¹, YANG Wen-yu¹, ZHENG Quan¹, CHE Xiao-yan². An easier method for generating recombinant adenovirus containing CD/TK fusi on gene driv-en by vascular endothelial growth factor promoter[J]. Journal of Southern Medical University, 2004, 24(01): 117-120.

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新法制备含VEGF启动子驱动的CD/TK融合基因重组腺病毒

An easier method for generating recombinant adenovirus containing CD/TK fusi on gene driv-en by vascular endothelial growth factor promoter

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