pTWIN质粒的改造及凋亡素组合肽基因工程菌的构建  

Construction of genetically engineered bacteria for apoptin combined peptides and modification of the pTWIN1 plasmid

作　　者：韩雪剑 闫佳敏 李鹤鸣 谭培珠 张慧姝 张丽秋 HAN Xue-jian;YAN Jia-min;LI He-ming;TAN Pei-zhu;ZHANG Hui-shu;ZHANG Li-qiu(Biotechnology Experimental Teaching Center,School of Basic Medical Sciences,Harbin Medical University,Harbin 150081,China)

机构地区：[1]哈尔滨医科大学基础医学院生物技术实验教学中心,黑龙江哈尔滨150081

出　　处：《哈尔滨医科大学学报》2021年第3期240-243,共4页Journal of Harbin Medical University

基　　金：黑龙江省卫生计生委科研课题(2017-160)。

摘　　要：目的构建凋亡素组合肽基因工程菌,改善组合肽的纯化工艺。方法以pTWIN1质粒为表达载体,构建凋亡素组合肽基因工程菌,对质粒进行改造,用6-His取代几丁质结合蛋白,实现用镍离子柱替换几丁质柱,一次层析纯化组合肽。结果构建了高水平表达凋亡素组合肽的基因工程菌;对pTWIN1质粒进行改造,用镍离子柱替代几丁质亲和柱纯化组合肽,实现了通过镍离子柱一次层析直接获得组合肽,内含肽自动断裂,不需要酶切,改造后的pTWIN1载体适用于凋亡素组合肽的开发,建立了纯化重组凋亡素组合肽方法。结论成功构建了高水平表达凋亡素组合肽的基因工程菌,成功地对pTWIN1质粒进行改造,为在生产中用镍离子柱纯化组合肽降低成本奠定了基础。Objective To construct a genetically engineered bacteria for apoptin combined peptides and improve the purification process of combined peptides. Methods The pTWIN1 plasmid was used as the expression vector to construct a genetically engineered apoptin combination peptide, and the plasmid was modified to replace the chitin binding protein with 6-His to realize the replacement of the chitin column with the nickel ion column and the purification of the combined peptide by one-time chromatography. Results A genetically engineered strain expressing high-level apoptin combination peptides was constructed;the pTWIN1 plasmid was modified, and a nickel ion column was used instead of a chitin affinity column to purify the combination peptides, and the combination peptides were directly obtained through a nickel ion column chromatography. The intein was automatically broken and did not require restriction enzyme digestion. The modified pTWIN1 vector was suitable for the development of apoptin combination peptides. A method for purifying recombinant apoptin combination peptides was established. Conclusion A genetically engineered strain expressing apoptin combination peptides at a high level is successfully constructed, and the pTWIN1 plasmid is successfully modified. The combined peptides lay the foundation for the use of nickel ion columns to purify the combined peptides in production to reduce costs.

关 键 词：凋亡素 凋亡素组合肽 基因工程 亲和层析 

分 类 号：R392.11[医药卫生—免疫学]