Objective
THE STUDY AND COMPARISON OF BOTH ENDOGENOUS AND ARTIFICIALLY CONSTRUCTED PLASMIDS IN DIFFERENT YEAST SPECIES IS OF GREAT IMPORTANCE TO UNDERSTAND THE ROLE AND FUNCTIONING OF THESE EXTRACHROMOSAL DNA'S. THE RESULTS OBTAINED FROM THIS STUDY COULD FILL THE EXISTING GAP BETWEEN ON ONE HAND THE DETAILED KNOWLEDGE ABOUT (ECONOMICALLY LESS INTERESTING) LABORATORY STRAINS AND ON THE OTHER HAND THE (ALMOST COMPLETE) ABSENCE OF SUCH KNOWLEDGE FOR INDUSTRIALLY IMPORTANT YEAST SPECIES.
THE DEVELOPMENT OF MORE STABLE VECTOR SYSTEMS FOR THE INDUSTRIAL WILL OPEN THE POSSIBILITY OF IMPROVING IMPORTANT CHARACTERISTICS IN THESE SPECIES AND THEIR WIDESPREAD USE IN, FOR EXAMPLE, THE AGRO-FOOD INDUSTRIES, STARCH DEGRADATION AND WASTE TREATMENT.
The genus Kluyveromyces is important in dairy industries and lately the pKD1 plasmid as well as the KARS based vectors have proved to be promising instruments in the physiological studies and genetic engineering of dairy yeasts.
Efficient expression vectors have been developed for the yeasts of industrial importance, in particular, the dairy yeasts.
Research has involved:
structural and functional characterization of the plasmid pKD1 and its derivative vectors;
analysis of the mechanism that gives autonomy and stability to the vectors derived from 2 and pKD1;
study of physiological regulation in Kluyveromyces lactis as a production organism.
The molecular structures of the plasmid pKD1 and of the autonomously replicating KARS sequence from K lactis chromosome were established. Plasmid and host functions important for the stable maintenance of the yeast plasmid were examined. Highly stable forms of pKD1 derived vectors were obtained. The host range of pKD1 was extended to several other species of Kluyveromyces. Shuttle deoxyribonucleic acid (DNA) banks were constructed for Kluyveromyces and Saccharomyces species. By combining the K lactis killer toxin gene sequence and pKD1 derived vectors, performing secretion vectors were obtained. Human interleukin-1 beta could be produced at high levels. To better control the gene expression of K lactis, several promoters were studied; in particular, new knowledge was obtained on the regulation of lactose metabolism and in the alcohol dehydrogenase system.
THE AIM OF THE RESEARCH IS TO DEVELOP MORE STABLE VECTOR SYSTEMS FOR TRANSFORMATION OF INDUSTRIAL YEAST SPECIES OTHER THAN S. CEREVISIAE. ENDOGENOUS AND ARTIFICIALLY CONSTRUCTED PLASMIDS WILL BE STUDIED FOR THEIR MAINTENANCE AND REPLICATION IN DIFFERENT SPECIES. IN PARTICULAR WILL BE STUDIED :
- THE ROLE OF ARS-SEQUENCES;
- THE EXISTENCE OF HOST-SPECIFIC STB BINDING PROTEIN;
- THE ROLE OF REP-SEQUENCES (ESPECIALLY WHEN UNDER CONTROL OF SPECIES SPECIFIC PROMOTORS).
THE INFORMATION SHOULD LEAD TO THE CONSTRUCTION OF A PLASMID THAT IS ABLE TO REPLICATE AND TRANSFORM S. CEREVISIAE, HANSENULA AND SCHWANNIOMYCES. AT THE SAME TIME, MUTANTS OF THESE SPECIES WILL BE OBTAINED IN ORDER TO TEST COMPLEMENTING GENES. PICHIA SPECIES WILL ALSO BE STUDIED AS AN INTERESTING SOURCE FOR SEVERAL GENES, FOR EXAMPLE THOSE INVOLVED IN XYLOSE FERMENTATION.
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CSC - Cost-sharing contractsCoordinator
40225 DUESSELDORF
Germany