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. 2015 Feb 4;15(1):17. doi: 10.1186/s12866-015-0354-3

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© Bruckbauer et al.; licensee BioMed Central. 2015

This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly credited. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.

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Mini-Tn 5/7 - lux aided promoter identification and capture in A. baumannii . The promoter-less mini-Tn5/7-lux element from pTn5/7LuxK5 was transposed into the A. baumannii strain ATCC19606 chromosome. Chromosomal DNA from a Km^r and luminescent exconjugant (IFD3) was isolated, digested with Acc65I, religated and transformed into E. coli pir-116 ⁺ strain MaH1. DNA from a Km^r transformants was sequenced with the luxC-specific primer P2385. The mini-Tn5/7-lux transposon was inserted in a gene annotated as A1S_0947 in strain ATCC17978 and the upstream A1S_0944-A1S_0945 intergenic region containing several possible promoters was PCR-amplified and cloned on a 329-bp StuI-DraIII fragment into pTn5/7LuxK5. The mini-Tn7-P _H1A-lux element was transposed into the attTn7 site on the ATCC19606 chromosome resulting in strain IFD5. Ten μl samples of an overnight culture of the indicated strains were spotted on an LB plate, grown overnight at 37°C and light emission was measured using a Xenogen IVIS imager.