A direct-selection vector derived from pColE3-CA38 and adapted for foreign gene expression.
The construction of a plasmid vector, pVT25, which allows an efficient and direct selection for transformed cells carrying recombinant plasmids is described. In this vector, the replicon and ApR gene from plasmid pBR327 are fused to the colE3 gene of pColE3-CA38, whereby positive selection is based on the inactivation of the lethal colicin E3 by the insertion of a foreign DNA fragment. However, pVT25 can be maintained within the Escherichia coli cells when complemented with another plasmid, pVT26, which expresses the colicin E3 immunity (imm) and the TcR phenotypes. Furthermore, pVT25 was used to regulate the expression of the synthetic human proinsulin gene fused to the colE3 gene at the single ClaI site. The production of the characteristic C-peptide of proinsulin, monitored by radioimmunoassay, was shown to be under the control of the inducible promoter of the colE3 gene.
Vernet
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Lau
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PC
Narang
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SA
Visentin
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LP
D001426:Bacterial Proteins / Q000235:genetics; D003086:Bacteriocin Plasmids; D003001:Cloning, Molecular; D003087:Colicins / Q000235:genetics; D004262:DNA Restriction Enzymes; D004269:DNA, Bacterial / Q000235:genetics; D004274:DNA, Recombinant; D015252:Deoxyribonucleases, Type II Site-Specific; D004926:Escherichia coli / Q000235:genetics; D029968:Escherichia coli Proteins; D005786:Gene Expression Regulation; D005798:Genes, Bacterial; D005813:Genes, Synthetic; D005822:Genetic Vectors; D006801:Humans; D010957:Plasmids; D011384:Proinsulin / Q000235:genetics
D001426:Bacterial Proteins; D003087:Colicins; D004269:DNA, Bacterial; D004274:DNA, Recombinant; D029968:Escherichia coli Proteins; C488855:ceiC protein, E coli; C007104:colicin immunity proteins; D011384:Proinsulin; D004262:DNA Restriction Enzymes; C032824:ATCGAT-specific type II deoxyribonucleases; D015252:Deoxyribonucleases, Type II Site-Specific