A new method for protein coexpression in Escherichia coli using two   incompatible plasmids

doi:10.1006/prep.2001.1453

CORC > 北京大学 > 生命科学学院

	A new method for protein coexpression in Escherichia coli using two incompatible plasmids
	Yang, W ; Zhang, L ; Lu, ZG ; Tao, W ; Zhao, ZH
刊名	蛋白质表达与纯化
	2001
关键词	CASPASE-ACTIVATED DNASE HIGH-LEVEL EXPRESSION CYTOCHROME-C APOPTOSIS INHIBITOR NUCLEASE FRAGMENTATION SOLUBILITY ICAD
DOI	10.1006/prep.2001.1453
英文摘要	It is commonly believed that incompatible plasmids carrying the same replicon cannot coexist stably in one Escherichia coli cell. However, we found that two incompatible plasmids carrying different antibiotic resistance genes, if under the selection pressure of the two antibiotics, can coexist in E. coli for at least 14 h, which is adequate for routine culture and protein expression. Based on this discovery, we developed a new method to coexpress foreign proteins in E. coli using two incompatible plasmids. The coding regions of the two subunits (DFF45 and DFF40) of the human DNA fragmentation factor (DFF) were cloned into two incompatible bacterial expression vectors-pET-21a with ampicillin resistance and pET-28a with kanamycin resistance, respectively. The two resulting plasmids were used to cotransform E. coli BL21(DE3) cells. After selection by ampicillin and kanamycin simultaneously, cotransformants that contain both recombinant plasmids were obtained. Induced by isopropyl beta -D-thiogalactoside, DFF45, and DFF40 were coexpressed efficiently in the presence of the two antibiotics. The coexpression product contained adequate soluble portions for both DFF45 and DFF40, while all DFF40 was insoluble if expressed alone. The coexpression product also exhibited the same caspase-activated DNase activity as its natural counterparts, which cannot be obtained if its two subunits are expressed separately. (C) 2001 Academic Press.; Biochemical Research Methods; Biochemistry & Molecular Biology; Biotechnology & Applied Microbiology; SCI(E); PubMed; 26; ARTICLE; 3; 472-478; 22
语种	英语
内容类型	期刊论文
源URL	[http://ir.pku.edu.cn/handle/20.500.11897/200554]
专题	生命科学学院
推荐引用方式 GB/T 7714	Yang, W,Zhang, L,Lu, ZG,et al. A new method for protein coexpression in Escherichia coli using two incompatible plasmids[J]. 蛋白质表达与纯化,2001.
APA	Yang, W,Zhang, L,Lu, ZG,Tao, W,&Zhao, ZH.(2001).A new method for protein coexpression in Escherichia coli using two incompatible plasmids.蛋白质表达与纯化.
MLA	Yang, W,et al."A new method for protein coexpression in Escherichia coli using two incompatible plasmids".蛋白质表达与纯化 (2001).