STRATEGY BASED ON PLASMID INCOMPATIBILITY FOR TN5 MUTAGENESIS IN ERWINIA-HERBICOLA ATCC-21998

Erwinia herbicola and Gluconobacter oxydans are ketogenic bacteria cap able of oxidation of hexose sugars to a number of keto-derivatives med iated by a consortium of enzymes known as dehydrogenases. In our studi es with G. oxydans, Tn5 transposition give rise to block mutants in th e direct glucose oxidation pathway. Some of these mutants have commerc ial value. But similar mutagenesis was not successful in E. herbicola because pSUP2021, the Tn5 carrying vector, stably replicated in this o rganism when transformed with the vector. Here we report a strategy fo r transposon mutagenesis in E. herbicola based on plasmid incompatibil ity to convert the stably maintaining plasmid pSUP2021 into a suicidal vector in presence of another plasmid pBR322::Tn5 incompatible with i t. The resultant transpogenesis generated auxotrophic mutants (for arg inine and lysine) in this commercially valuable organism. The study al so confirmed that the native plasmid (pVQ1) harboured by E. herbicola has its origin non-homologous to that of pMB1.