Inducible amplification of gene copy number and heterologous protein production in the yeast Kluyveromyces lactis.
Heterologous protein production can be doubled by increasing the copy number of the corresponding heterologous gene. We constructed a host-vector system in the yeast Kluyveromyces lactis that was able to induce copy number amplification of pKD1 plasmid-based vectors upon expression of an integrated copy of the plasmid recombinase gene. We increased the production and secretion of two heterologous proteins, glucoamylase from the yeast Arxula adeninivorans and mammalian interleukin-1beta, following gene dosage amplification when the heterologous genes were carried by pKD1-based vectors. The choice of the promoters for expression of the integrated recombinase gene and of the episomal heterologous genes are critical for the mitotic stability of the host-vector system.
Morlino
G B
GB
Department of Cell and Developmental Biology, University of Rome "La Sapienza," Rome 00185, Italy.
Tizzani
L
L
Fleer
R
R
Frontali
L
L
Bianchi
M M
MM
D000818:Animals; D003001:Cloning, Molecular / Q000379:methods; D004254:DNA Nucleotidyltransferases / Q000235:genetics*; D005784:Gene Amplification; D005822:Genetic Vectors; D005087:Glucan 1,4-alpha-Glucosidase / Q000096:biosynthesis / Q000235:genetics; D019426:Integrases; D007375:Interleukin-1 / Q000096:biosynthesis* / Q000235:genetics*; D007716:Kluyveromyces / Q000201:enzymology / Q000235:genetics*; D008322:Mammals; D010957:Plasmids; D016133:Polymerase Chain Reaction; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D045522:Recombinases; D015003:Yeasts / Q000235:genetics
D007375:Interleukin-1; D011993:Recombinant Fusion Proteins; D045522:Recombinases; D004254:DNA Nucleotidyltransferases; D019426:Integrases; C115006:integron integrase IntI1; D005087:Glucan 1,4-alpha-Glucosidase