膜联蛋白32与低分子质量单链尿激酶融合基因的构建及高效表达  

Construction and expression of a low relative molecular mass single-chain urokinase-type plasminogen activator(ScuPA32k) and annexin32

作　　者：颜宏利[1] 孙树汉[1] 陈蕊雯[1] 杨湘越[1] 

机构地区：[1]第二军医大学基础医学部医学遗传学教研室,上海200433

出　　处：《第二军医大学学报》2002年第4期378-380,共3页Academic Journal of Second Military Medical University

基　　金：国家"863"计划资助项目(2001AA213111).

摘　　要：目的：构建膜联蛋白32（annexin32，Anx32）和低相对分子质量单链尿激酶（ScuPA32k）的融合基因，并在大肠杆菌系统内作表达。方法：利用重叠区扩增法进行基因拼接，然后在谷胱甘肽S转移酶原核表达系统内表达，Western印迹验证表达产物。结果：重叠区扩增得到 1．8 kb融合基因，该基因在原核表达系统内得到高效表达，表达量占菌体总蛋白的 26％，Western印迹证明表达产物正确。结论：用重叠区扩增法进行基因拼接快速、简便。Objective: To obtain a chimeras composed of annexin32 (Anx32) and low relative molecular mass of single-chain urokinaseutype plasminogen activator(ScuPA32k). Methods: A fusion gene Anx32-ScuPA32k was constructed by over-lap extension gene splicing and then inserted into plasmid pGEX-5T. The recombined plasmid was transformed into E. colistrain K802 and induced by IPTG. Western blotting was used to identify the products. Results: A 1. 8 kb gene fragment wasobtained by gene splicing. After inserted into the expression plasmid,a high level recombinant was induced by 0. 5 mol/LIPTG. Western Blotting indicated that the recombinant was able to react with anti-sera against annexin 32. Conclusion: Genesplicing by overlap extension is a fast and easy way to splice 2 genes.

关 键 词：膜联蛋白32 低相对分子质量单链尿激酶 基因拼接 基因表达 重叠区扩增法 大肠杆菌 

分 类 号：R973.2[医药卫生—药品]