New selectable marker/auxotrophic host strain combinations for molecular genetic manipulation of Pichia pastoris.
We describe the isolation and characterization of three new biosynthetic genes-ARG4, ADE1, and URA3-from the methylotrophic yeast Pichia pastoris. The predicted products of the genes share significant sequence similarity to their Saccharomyces cerevisiae counterparts, namely argininosuccinate lyase, PR-aminoimidazolesuccinocarboxamide synthase, and orotidine-5'-phosphate decarboxylase, respectively. Along with the previously described HIS4 gene, each gene was incorporated as the yeast selectable marker into a set of shuttle vectors designed to express foreign genes in P. pastoris. In addition, we have constructed a series of host strains containing all possible combinations of ade1, arg4, his4, and ura3 auxotrophies to be used with these new vectors.
Department of Biochemistry and Molecular Biology, Oregon Graduate Institute of Science and Technology, 20000 N.W. Walker Road, Beaverton, OR 97006-8921, USA.
G P
GP
Lin Cereghino
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Lin Cereghino
A J
AJ
Sunga
M A
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Johnson
M
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Lim
M A
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Gleeson
J M
JM
Cregg
D000595:Amino Acid Sequence; D001123:Argininosuccinate Lyase / Q000235:genetics; D003001:Cloning, Molecular; D004271:DNA, Fungal / Q000737:chemistry / Q000235:genetics; D005656:Fungal Proteins / Q000235:genetics* / Q000378:metabolism; D005800:Genes, Fungal / Q000235:genetics; D005816:Genetic Complementation Test; D005819:Genetic Markers; D000432:Methanol / Q000378:metabolism; D008969:Molecular Sequence Data; D009154:Mutation; D009964:Orotidine-5'-Phosphate Decarboxylase / Q000235:genetics; D010453:Peptide Synthases / Q000235:genetics; D010843:Pichia / Q000201:enzymology / Q000235:genetics*; D010957:Plasmids / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid
D004271:DNA, Fungal; D005656:Fungal Proteins; D005819:Genetic Markers; D011993:Recombinant Fusion Proteins; D009964:Orotidine-5'-Phosphate Decarboxylase; D001123:Argininosuccinate Lyase; D010453:Peptide Synthases; C022379:phosphoribosylaminoimidazole-succinocarboxamide synthetase; D000432:Methanol