Integrative transformation by homologous recombination in the zygomycete Mucor circinelloides

Abstract

Transformation of a Mucor circinelloides Leu- strain with the plasmid pAD45, harbouring the wild-type allele (leuA+) and a chymosin gene, led to the identification of mitotically stable transformants after one to three vegetative growth cycles on non-selective medium. Southern analysis of the stable transformed strains demonstrated that the vector is integrated, as an intact molecule, into the resident Mucor leuA locus. Retransformation of Escherichia coli with genomic DNA restricted with enzymes having no or only a single recognition site within the inserted sequence did not permit isolation of plasmids or fragments carrying the leuA or chymosin gene.