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. 2004 Jul;167(3):1069–1077. doi: 10.1534/genetics.104.026682

Insertions of mini-Tn10 transposon T-POP in Salmonella enterica sv. typhi.

Alejandro A Hidalgo 1, A Nicole Trombert 1, J C Castro-Alonso 1, Carlos A Santiviago 1, Bruno R Tesser 1, Philip Youderian 1, Guido C Mora 1
PMCID: PMC1470937  PMID: 15280224

Abstract

We have mutagenized a clinical strain of Salmonella enterica sv. typhi with mini-transposon Tn10dTet (T-POP) to obtain conditional lethal (tetracycline-dependent) mutants with T-POP insertions upstream of essential genes. Generalized transducing phage P22 was used to introduce T-POP from a S. typhimurium donor into a S. typhi recipient. Chromosomal DNA was purified from the mutagenized donor strains, fragmented, and then electroporated into S. typhi to backcross the original T-POP insertions. Four tetracycline-dependent mutants with two distinct terminal phenotypes were found among 1700 mutants with T-POP insertions. When grown in the absence of tetracycline, two of the four tetracycline-dependent mutants arrest at a late stage in the cell cycle, can be rescued by outgrowth in media with tetracycline, and define a reversible checkpoint late in the cell cycle. One of these insertions creates an operon fusion with a gene, yqgF, that is conserved among gram-negative bacteria and likely encodes an essential Holliday junction resolvase. T-POP insertions can be used not only to identify essential S. typhi genes but also to reveal novel phenotypes resulting from the depletion of their products.

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Selected References

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