A novel system of genetic transformation allows multiple integrations of a desired gene in Saccharomyces cerevisiae chromosomes

作者：Guerra Odanir Garcia; Rubio Ileana G S; da Silva Filho Claudionor Gomes; Bertoni Regiane Aparecida; dos Santos Govea Rute Cardoso; Vicente Elisabete Jose^*

来源：Journal of Microbiological Methods, 2006, 67(3): 437-445.

DOI：10.1016/j.mimet.2006.04.014

• glucoamylase
• retrotransposon Ty1
• S. cerevisiae transformation
• yeast linear transformation vector
• yeast transformation vector without selective marker
• delta integration

Increasing industrial competitiveness and productivity demand that recombinant yeast strains, used in many different processes, be constantly adapted and/or genetically improved to suit changing requirements. Among yeasts, Saccharomyces cerevisiae is the best-studied organism, and the most frequently employed yeast in industrial processes. In the present study, laboratory strains and industrial S. cerevisiae strains were stably transformed with a novel vector containing the glucoamylase cDNA of Aspergillus awamori flanked by delta-sequences (delta Gluco delta), and lacking a positive selection marker. Co-transformation with known plasmids allowed selection by auxotrophic complementation of the leu2 mutation and/or geneticin resistance (G418(R)). In all cases, several copies of the delta Gluco delta vector were inserted into the genome of the yeast cell without selective pressure, showing 100% stability after 80 generations. Transformation frequency of the new vector was similar for S. cerevisiae laboratory strains and industrial wild-type S. cerevisiae strains. This novel genetic transformation system is versatile and suitable to introduce several stable copies of a desired expression cassette into the genome of different S. cerevisiae yeast strains.

• 出版日期2006-12