Construction of a recF deletion mutant of Azotobacter vinelandii and its characterization

A deletion was engineered in the cloned recF gene by digestion with suitabl e restriction endonucleases and a tetracycline resistance gene cartridge wa s inserted. The mutation was subsequently transferred to the Azotobacter vi nelandii chromosome by double cross-over under pressure of tetracycline sel ection. A recF recA mutant was also constructed in a similar manner. The mu tations were found to be stable and mutation of the wild-type recF gene was confirmed by Southern blot hybridization. Both the mutants were UV sensiti ve and recombination deficient. Mutations in genes involved in nitrogen fix ation in A. vinelandii are rather frequent and obtained comparatively easil y despite of the presence of multiple identical chromosomes in A. vinelandi i. It has been speculated that some kind of 'homogenotization' process oper ates which is responsible for the 'transmission' of mutation from one chrom osome to all the chromosomes. This process is not affected by a mutation in recF or recA or in both recF and recA. (C) 1999 Federation of European Mic robiological Societies. Published by Elsevier Science B.V. All rights reser ved.