doiSerbia

Effect of the kanamycin resistance marker on stability of 2μ-based expression plasmids

Stanković Nada (Institut za molekularnu genetiku i genetičko inženjerstvo, Beograd)
Vasiljević Branka (Institut za molekularnu genetiku i genetičko inženjerstvo, Beograd)
Ljubijankić Goran (Institut za molekularnu genetiku i genetičko inženjerstvo, Beograd)

In this paper we describe the effect of the kanamycin resistance gene (Kmr) on 2μm-based plasmid maintenance in Saccharomyces cerevisiae. The influence of this marker gene on the loss of the stable model-vectors proved to be constant, as well as independent of carbon source and culture growth rates. In strains for GALUAS - driven heterologous protein production introduction of Kmr resulted in curing of the yeast episomal plasmid (YEp) from the population in a small number of generations. Application of selective pressure on the strain producing recombinant penicillin G amidase (rPGA) did not provide the expected increase of protein yield. The influence of genetic elements for heterologous protein production on vector stability was examined, and the most destabilizing factors prove to be the presence and expression of the foreign gene.

Keywords: 2μm plasmid, kanamycin resistance, penicillin G amidase, plasmid stability, Saccharomyces cerevisiae, selective markers