Validation of a Tn5 transposon mutagenesis system for Gluconacetobacter diazotrophicus through characterization of a flagellar mutant View Full Text


Ontology type: schema:ScholarlyArticle     


Article Info

DATE

2008-04

AUTHORS

Luc F. M. Rouws, Jean L. Simões-Araújo, Adriana S. Hemerly, José I. Baldani

ABSTRACT

Gluconacetobacter diazotrophicus is a nitrogen-fixing bacterium, which was originally isolated from the interior of sugarcane plants. The genome of strain PAL5 of G. diazotrophicus has been completely sequenced and a next step is the functional characterization of its genes. The aim of this study was to establish an efficient mutagenesis method, using the commercial Tn5 transposon EZ::Tn5Tnp Transposome (Epicentre). Up to 1 x 10(6) mutants per microgram of transposome were generated in a single electroporation experiment. Insertion-site flanking sequences were amplified by inverse PCR and sequenced for 31 mutants. For ten of these mutants, both insertion flanks could be identified, confirming the 9 bp duplication that is typical for Tn5 transposition. Insertions occurred in a random fashion and were genetically stable for at least 50 generations. One mutant had an insertion in a homolog of the flagellar gene flgA, and was therefore predicted to be affected in flagella-dependent traits and used to validate the applied mutagenesis methodology. This mutant lacked flagella and was non-motile on soft agar. Interestingly, it was also strongly affected in the ability to form biofilm on glass wool. More... »

PAGES

397-405

References to SciGraph publications

Identifiers

URI

http://scigraph.springernature.com/pub.10.1007/s00203-007-0330-x

DOI

http://dx.doi.org/10.1007/s00203-007-0330-x

DIMENSIONS

https://app.dimensions.ai/details/publication/pub.1017750064

PUBMED

https://www.ncbi.nlm.nih.gov/pubmed/18060666


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