ColE1-compatible vectors for high-level expression of cloned DNAs from the T7 promoter.
A new family of T7-based expression plasmids with unique features is described. The plasmid origin of replication (ori), derived from P15A, is compatible with that of ColE1-derived plasmids, which facilitates the co-production of proteins from these vectors and from ColE1-derived T7 expression vectors in the same cell. The plasmids are medium-copy-number and also carry the M13 ori. Consequently, both double- and single-stranded DNA can be easily obtained. The plasmids encode KmR, thus avoiding the potential for plasmid loss associated with ApR-based systems. One of the plasmids carries the lacI gene, to allow for more stringent regulation of the production of potentially toxic proteins. When the plasmids are introduced into an Escherichia coli strain such as BL21(DE3), which contains the T7 polymerase-encoding gene under control of the lacUV5 promoter, addition of IPTG initiates the production of high levels of the recombinant protein.
Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, CT 06520.
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M
Munson
P F
PF
Predki
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L
Regan
D003086:Bacteriocin Plasmids; D017123:Bacteriophage T7 / Q000235:genetics*; D001483:Base Sequence; D003001:Cloning, Molecular; D004274:DNA, Recombinant / Q000235:genetics*; D004277:DNA, Single-Stranded; D004926:Escherichia coli / Q000235:genetics; D005822:Genetic Vectors; D008969:Molecular Sequence Data; D010957:Plasmids; D011401:Promoter Regions, Genetic; D011994:Recombinant Proteins / Q000096:biosynthesis
D004274:DNA, Recombinant; D004277:DNA, Single-Stranded; D011994:Recombinant Proteins