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Date of Award
2002
Document Type
Thesis - Pacific Access Restricted
Degree Name
Master of Science (M.S.)
Department
Biological Sciences
First Advisor
Geoff Lin-Cereghino
First Committee Member
Craig A. Vierra
Second Committee Member
Lisa A. Wrischnik
Abstract
The methylotrophic yeast, Pichia pastoris, has been used as a protein expression system to express over 500 heterologous proteins. P. pastoris provides many advantages over other organisms that have been utilized for this purpose. In this project, we developed a new host/selectable marker and auxotrophic strains of P. pastoris based on methionine biosynthesis to increase P. pastoris's versatility as a host for homologous protein expression. This was accomplished by selecting for a yeast that is deficient in methionine biosynthesis, P. pastoris (yJC239), and gene complementation through transformation with a genomic DNA library.
Bioinformatics show that the P. pastor is MET gene has 54% amino acid identity with 68% similarity to the S. cerevisiae MET2 gene, which codes for homoserine O-transacetylase. We have constructed expression vectors for intracellular and extracellular expression of proteins with the MET2 marker and have also constructed strains with various auxotrophs including me/2.
Pages
61
Recommended Citation
Thor, Der. (2002). Cloning and characterization of MET2 in Pichia pastoris. University of the Pacific, Thesis - Pacific Access Restricted. https://scholarlycommons.pacific.edu/uop_etds/570
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