Identification of a Yarrowia lipolytica acetamidase and its use as a yeast genetic marker.
BACKGROUND
Yarrowia lipolytica is an oleaginous yeast that can be genetically engineered to produce lipid and non-lipid biochemicals from a variety of feedstocks. Metabolic engineering of this organism usually requires genetic markers in order to select for modified cells. The potential to combine multiple genetic manipulations depends on the availability of multiple or recyclable selectable markers.


RESULTS
We found that Y. lipolytica has the ability to utilize acetamide as the sole nitrogen source suggesting that the genome contains an acetamidase gene. Two potential Y. lipolytica acetamidase gene candidates were identified by homology to the A. nidulans acetamidase amdS. These genes were deleted in the wild-type Y. lipolytica strain YB-392, and deletion strains were evaluated for acetamide utilization. One deletion strain was unable to grow on acetamide and a putative acetamidase gene YlAMD1 was identified. Transformation of YlAMD1 followed by selection on acetamide media and counterselection on fluoroacetamide media showed that YlAMD1 can be used as a recyclable genetic marker in Saccharomyces cerevisiae and Ylamd1Δ Y. lipolytica.


CONCLUSIONS
These findings add to our understanding of Y. lipolytica nitrogen utilization and expand the set of genetic tools available for engineering this organism, as well as S. cerevisiae.
['Hamilton|Maureen|M|http://orcid.org/0000-0002-2995-724X', 'Consiglio|Andrew L|AL|', 'MacEwen|Kyle|K|', 'Shaw|A Joe|AJ|', 'Tsakraklides|Vasiliki|V|']
D000081:Acetamides D000581:Amidohydrolases D005819:Genetic Markers D060847:Metabolic Engineering D012441:Saccharomyces cerevisiae D014170:Transformation, Genetic D025062:Yarrowia
Yarrowia lipolytica Acetamidase Acetamide Fluoroacetamide Counterselectable marker Dominant marker Marker recycling Marker removal