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A tandem reporter plasmid carrying promoterless xyIE and gfpmut3 genes downstream of the MCS. The plasmid can be conjugated into a range of bacterial strains difficult to transform by electroporation. pMC-Tandem is a low copy number plasmid.

Uses reporters XyIE and GFP to screen and identify transcriptional regulators and conditions which favour gene expression from different promotors that can be inserted in the MCS. Based on broad-host-range shuttle vector pMIDG100. XyIE allows for rapid screening of bacteria on agar plates due to development of yellow colour when sprayed with catechol.

Figure: Construction of the reporter vector, pMC-Tandem. (A) pMIDG100 (49) containing aphA3 (kanamycin resistance), gfpmut3 (GFP), and mob (mobilization protein). (B) pMIDG300 was derived from pMIDG100 by cloning the cat gene (chloramphenicol resistance) from S. aureus pC194 (30) into the two EcoRV sites, replacing the aphA3 gene. (C) pMC-Tandem was derived from pMIDG300 by insertion of the xylE gene (catechol 2,3-dioxygenase) from pJFF224-NX:xylE (18) into the unique BamHI and NheI sites upstream of gfpmut3. Relevant restriction sites are labeled as follows: A, ApaI; B, BamHI; EI, EcoRI; EV, EcoRV; K, KpnI; N, NheI; P, PstI; Xb, XbaI; Xh, XhoI. All labeled restriction sites are unique except for EcoRV, which cuts twice in pMIDG100.

 

To obtain pMC-Tandem, please contact the Quicktech team directly (quicktech@imperialinnovations.co.uk).

 

For shuttle vectors, please see pMC-Express and pMK-Express

 

 

Reference:

New Plasmid Tools for Genetic Analysis of Actinobacillus pleuropneumoniae and Other Pasteurellaceae
Janine T. BosséAndrew L. DurhamAndrew N. RycroftJ. Simon KrollPaul R. Langford

Internal case number: 8156

pMC-Tandem

A tandem reporter plasmid to screen and identify transcriptional regulators and conditions in Pasteurellaceae

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