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1986•FEMS Microbiology LettersOpen access

A novel plasmid vector allowing positive selection for cloned fragments

M Garrido

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Abstract

The use of plasmid pMM102 as a positive selection cloning vector is described. This plasmid is derived from pBR322 and contains the DNA encoding microcin B17 production and immunity. RecA−Escherichia coli K12 cells containing this plasmid are unable to grow in minimal medium. Inactivation of any of the 4 genes required for microcin production allows the bacterial host to produce colonies. This property has been used to clone DNA inserts in the unique sites for restriction endonucleases BglII, SacI, SacII and SmaI in pMM102. DNA fragments with asymmetrical termini of many different kinds can also be cloned. We have also identified a fragment in the wild-type plasmid pMccB17 that suppresses the pMM102-induced growth inhibition.

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The use of plasmid pMM102 as a positive selection cloning vector is described. This plasmid is derived from pBR322 and contains the DNA encoding microcin B17 production and immunity. RecA−Escherichia coli K12 cells containing this plasmid are unable to grow in minimal medium. Inactivation of any of the 4 genes required for microcin production allows the bacterial host to produce colonies. This property has been used to clone DNA inserts in the unique sites for restriction endonucleases BglII, SacI, SacII and SmaI in pMM102. DNA fragments with asymmetrical termini of many different kinds can also be cloned. We have also identified a fragment in the wild-type plasmid pMccB17 that suppresses the pMM102-induced growth inhibition.

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Available abstract

The use of plasmid pMM102 as a positive selection cloning vector is described. This plasmid is derived from pBR322 and contains the DNA encoding microcin B17 production and immunity. RecA−Escherichia coli K12 cells containing this plasmid are unable to grow in minimal medium. Inactivation of any of the 4 genes required for microcin production allows the bacterial host to produce colonies. This property has been used to clone DNA inserts in the unique sites for restriction endonucleases BglII, SacI, SacII and SmaI in pMM102. DNA fragments with asymmetrical termini of many different kinds can also be cloned. We have also identified a fragment in the wild-type plasmid pMccB17 that suppresses the pMM102-induced growth inhibition.

Key concepts: Plasmid, BglII, PBR322, Cloning vector, Biology, Escherichia coli, Plasmid preparation, Restriction enzyme

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