Fig. 8

Construction of Y. lipolytica strains overexpressing ylGAL genes. The auxotrophic PO1d strain was used as the acceptor strain. The genes were inserted one by one to create strains that overexpressed different combinations of the ylGAL genes; URA3ex and LEU2ex were used as selection markers. To recover prototrophy in some of the Y. lipolytica strains (gray squares), cassettes containing the URA3 excisable marker or the purified SalI fragment of a pINA62 plasmid that contained the LEU2 gene were inserted into the transformants’ genomes [28]. In turn, to recover URA3 and LEU2 auxotrophy, which was necessary to continue the transformation process, a JME547 plasmid containing Cre-Lox recombinase was used [35]