Multiple gene deletion of Mannheimia succiniciproducens using mutant lox sequences

Abstract

A markerless gene deletion method was developed to generate genetically modified Mannheimia succiniciproducens, which is a rumen bacterium producing succinic acid as a major metabolite. Cre recombinase was transiently expressed using a temperature sensitive plasmid to excise the antibiotic marker between the mutant lox sites. As a demonstration of the method, ldhA gene and oadGAB operon was sequentially deleted without leaving an antibiotic marker inside the genome of M. succiniciproducens.