AILV1 GENE FROM THE YEAST ARXULA-ADENINIVORANS LS3 - A NEW SELECTIVE TRANSFORMATION MARKER

The ILV1 gene of the yeast Arxula adeninivorans LS3 (AILV1) has been c loned from a genomic library, characterized and used as an auxotrophic selection marker for transformation of plasmids into this yeast. One copy of the gene is present in the Arxula genome, comprising 1653 bp a nd encoding 550 amino acids of the threonine deaminase. The protein se quence is similar (60.55%) to that of the threonine deaminase from Sac charomyces cerevisiae encoded by the gene ILV1. The protein is enzymat ically active during the whole period of cultivation, up to 70 h. Maxi mal activities, as well as protein concentrations of this enzyme, were achieved after cultivation times of 20-36 h. The AILV1 gene is a suit able auxotrophic selection marker in transformation experiments using an Arxula adeninivorans ilv1 mutant and a plasmid containing this gene , which is fused into the 25S rDNA of Arxula adeninivorans. One to thr ee copies of the linearized plasmid were integrated into the 25S rDNA by homologous recombination. Transformants resulting from complementat ion of the ilv1 mutation can be easily and reproducibly selected and i n addition are mitotically stable. Therefore, the described system is preferred to the conventional selection for hygromycin B resistance. ( C) 1998 John Wiley & Sons, Ltd.