Multicopy plasmids containing the gene for the transcriptional activator LAC9 are not tolerated by K. lactis cells.
The high copy number 2-microns DNA-like Kluyveromyces plasmid pKD1 was extremely unstable in Kluyveromyces lactis when carrying the gene for the regulatory protein LAC9, a transcriptional activator involved in the induction of the LAC and GAL genes. Transformants of a lac9 mutant strain normally contained rearranged plasmids and all were Lac-, indicating that the LAC9 gene was inactive. Lac+ "revertants" could be obtained from Lac- transformants by selection on lactose plates. In some of these, the pKD1-based plasmid was stably maintained by being integrated into the chromosome of the cell; in others, the disrupted chromosomal gene was restored by a gene conversion event. None of the Lac+ revertants had more than one intact LAC9 gene, an indication that LAC9 overexpression affects cell viability.
Breunig
K D
KD
Institut für Mikrobiologie, Universität Düsseldorf, Federal Republic of Germany.
D015139:Blotting, Southern; D004268:DNA-Binding Proteins; D005656:Fungal Proteins / Q000235:genetics*; D005796:Genes; D005800:Genes, Fungal; D005822:Genetic Vectors; D007716:Kluyveromyces / Q000235:genetics*; D010957:Plasmids; D015183:Restriction Mapping; D004718:Saccharomycetales / Q000235:genetics*; D014157:Transcription Factors / Q000235:genetics*
D004268:DNA-Binding Proteins; D005656:Fungal Proteins; C058596:LAC9 protein, Kluyveromyces lactis; D014157:Transcription Factors