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Figure 1 | BMC Microbiology

Figure 1

From: Fast, easy and efficient: site-specific insertion of transgenes into Enterobacterial chromosomes using Tn7 without need for selection of the insertion event

Figure 1

Transposition vector. The vector pGRG25 contains a multiple cloning site (MCS) with unique restriction sites for Avr II, Not I, Pac I, and Xho I, flanked by the terminal repeats of Tn7. Cloning of the gene of interest into these sites, in between the Tn7 arms, will result in a plasmid that can be used to insert the transgene into the chromosome. araC is the regulator of the arabinose-inducible promoter PBAD (though in all experiments presented, leaky expression was sufficient). tnsABCD are the genes required for transposition of Tn7. rrnD is the transcription terminator. bla encodes β-lactamase which allows selection of ampicillin resistance when transforming the plasmid into bacteria. oriT is the RP4 conjugal transfer origin. pSC101 ori ts is the temperature sensitive origin of replication which functions at 32°C, but not at 42°C. Vector pGRG36 is identical except for the addition of a Sma I site to the MCS. The sequence of pGRG25 and pGRG36 can be accessed at GenBank (accession # DQ460223).

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BMC Microbiology

ISSN: 1471-2180

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