Strategy based on plasmid incompatibility for Tn5 mutagenesis in Erwinia herbicola ATCC 21998

摘要

Erwinia herbicola and Gluconobacter oxydans are ketogenic bacteria capable of oxidation of hexose sugars to a number of keto-derivatives mediated by a consortium of enzymes known as dehydrogenases. In our studies with G. oxydans, Tn5 transposition give rise to block mutants in the direct glucose oxidation pathway. Some of these mutants have commercial value. But similar mutagenesis was not successful in E. herbicola because pSUP2021, the Tn5 carrying vector, stably replicated in this organism when transformed with the vector. Here we report a strategy for transposon mutagenesis in E. herbicola based on plasmid incompatibility to convert the stably maintaining plasmid pSUP2021 into a suicidal vector in presence of another plasmid pBR322::Tn5 incompatible with it. The resultant transpogenesis generated auxotrophic mutants (for arginine and lysine) in this commercially valuable organism. The study also confirmed that the native plasmid (pVQ1) harboured by E. herbicola has its origin non-homologous to that of pMB1. [References: 16]