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2012

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Si us plau utilitzeu sempre aquest identificador per citar o enllaçar aquest document: https://hdl.handle.net/2445/32172

A chromosomal insertion toolbox for promoter probing, mutant complementation and pathogenicity studies in Ralstonia solanacearum

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Recurs relacionat

http://dx.doi.org/10.1094/MPMI-07-11-0201

Resum

We describe here the construction of a delivery system for stable and directed insertion of gene constructs in a permissive chromosomal site of the bacterial wilt pathogen Ralstonia solanacearum. The system consists of a collection of suicide vectors the Ralstonia chromosome (pRC) series that carry an integration element flanked by transcription terminators and two sequences of homology to the chromosome of strain GMI1000, where the integration element is inserted through a double recombination event. Unique restriction enzyme sites and a GATEWAY cassette enable cloning of any promoter::gene combination in the integration element. Variants endowed with different selectable antibiotic resistance genes and promoter::gene combinations are described. We show that the system can be readily used in GMI1000 and adapted to other R. solanacearum strains using an accessory plasmid. We prove that the pRC system can be employed to complement a deletion mutation with a single copy of the native gene, and to measure transcription of selected promoters in monocopy both in vitro and in planta. Finally, the system has been used to purify and study secretion type III effectors. These novel genetic tools will be particularly useful for the construction of recombinant bacteria that maintain inserted genes or reporter fusions in competitive situations (i.e., during plant infection).

Matèries

Cromosomes, Genètica, Bacteris patògens, Patologia vegetal

Matèries (anglès)

Chromosomes, Genetics, Pathogenic bacteria, Plant pathology

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Articles publicats en revistes (Genètica, Microbiologia i Estadística)

Citació

MONTEIRO, Freddy, et al. A chromosomal insertion toolbox for promoter probing, mutant complementation and pathogenicity studies in Ralstonia solanacearum. Molecular Plant-Microbe Interactions. 2011. Vol. 25, num. 4, pags. 557-568. ISSN 0894-0282. [consulted: 16 of August of 2026]. Available at: https://hdl.handle.net/2445/32172

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