Improved shuttle vectors for cloning and high-level Cu(2+)-mediated expression of foreign genes in yeast.
New yeast episomal vectors having a high degree of utility for cloning and expression in Saccharomyces cerevisiae are described. One vector, pYEULlacZ, is based on pUC19 and employs the pUC19 multiple cloning site for the selection of recombinants in Escherichia coli by lacZ inactivation. In addition, the vector contains two genes, URA3 and leu2-d, for selection of the plasmid in ura3 or leu2 yeast strains. The presence of the leu2-d gene appears to promote replication at high copy numbers. The introduction of CUP1 cassettes allows these plasmids to direct Cu(2+)-regulated production of foreign proteins in yeast. We show the production of a helminth antigen as an example of the vector application.
CSIRO Division of Biomolecular Engineering, Parkville, Victoria, Australia.
I G
IG
Macreadie
O
O
Horaitis
A J
AJ
Verkuylen
K W
KW
Savin
D000818:Animals; D000947:Antigens, Helminth / Q000032:analysis / Q000235:genetics*; D001483:Base Sequence; D003001:Cloning, Molecular / Q000187:drug effects / Q000379:methods*; D003300:Copper / Q000494:pharmacology*; D004926:Escherichia coli / Q000235:genetics*; D005798:Genes, Bacterial; D005800:Genes, Fungal; D005822:Genetic Vectors; D006639:Histidine / Q000378:metabolism; D008969:Molecular Sequence Data; D008970:Molecular Weight; D015345:Oligonucleotide Probes; D010957:Plasmids; D011994:Recombinant Proteins / Q000032:analysis; D015183:Restriction Mapping; D012441:Saccharomyces cerevisiae / Q000187:drug effects / Q000235:genetics*; D014170:Transformation, Genetic; D014254:Trichostrongylus / Q000235:genetics; D014364:Tryptophan / Q000378:metabolism
D000947:Antigens, Helminth; D015345:Oligonucleotide Probes; D011994:Recombinant Proteins; D006639:Histidine; D003300:Copper; D014364:Tryptophan