Development of a Yeast Strain that Secretes β-galactosidase

Katy Harris-Forkner Saint Mary's College, Lauren Johnson Saint Mary's College
Faculty Sponsor(s): Don Paetkau Saint Mary's College
Bacterial and yeast whole-organism biosensors have been created using a number of reporter genes including LacZ, which encodes the β-galactosidase protein. LacZ works particularly well in bacteria because its protein is secreted. The secretion creates a vibrant blue color that is easy to detect. Yeast cells make hardier biosensors than bacteria, but they do not naturally secrete the β-galactosidase protein, making the color harder to detect. PCR was used to successfully amplify the 3074 kb LacZ gene from a bacterial plasmid (pCM176). Recombination/integration sequences were added to LacZ gene during this step and were successfully used to insert the PCR fragment into the pD1214-AA yeast expression plasmid. A portion of the TEF-αAmylase-ORF was used to control secretion of the β-galactosidase protein from this plasmid. The new vector was inserted into the Y187 Saccharomyces cerevisiae cells. Secretion was tested by growth on minus uracil standard dropout media containing X-gal. Further enhancement of the secretion process is proposed using a using yeast mutagenesis screen. Secretion of other proteins from yeast, such as the cellulase gene from symbiotes found in the termite gut, is also being studied.
Biochemistry & Molecular Biology
Poster Presentation

When & Where

Irwin Library 3rd Floor