CONSTRUCTION OF AN AMYLOLYTIC YEAST BY MULTIPLE INTEGRATION OF THE ASPERGILLUS-AWAMORI GLUCOAMYLASE GENE INTO A SACCHAROMYCES-CERVISIAE CHROMOSOME

A plasmid containing the glucoamylase cDNA gene (glu) from Aspergillus awamori, a portion of Saccharomyces cerevisiae ribosomal DNA, S. cere visiae urd gene deficient in promoter, and bacterial cloning vector pS P72 was constructed for the integration of an A. awamori glu gene into the genome of S. cerevisiae. Several transformants were able to form halos around colonies on the plate containing 2% soluble starch. This indicated the secretion of glucoamylase by the yeasts. Glucoamylase pr oduction by transformant G23-8 reached 162 U l(-1). Southern hybridiza tion analysis revealed that the plasmid was integrated into the host c hromosome in tandem repeat and dispersion copies. It was estimated tha t the integrants carried up to 140 glu gene copies per genome. The int egrants were mitotically stable for 50 generations without selective p ressure. (C) 1998 Elsevier Science Inc.