1 Department of Micro- and Nanotechnology, Technical University of Denmark2 Department of Systems Biology, Technical University of Denmark3 Eucaryotic Molecular Cell Biology, Department of Systems Biology, Technical University of Denmark4 Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark5 Projects, Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark6 Eukaryotic Molecular Cell Biology, Department of Biotechnology and Biomedicine, Technical University of Denmark7 Novo Nordisk A/S
When studying protein folding and secretion the general conception is that all cells in a population express an equal amount of protein. Recent work has shown that expression levels vary greatly in cell populations which express proteins on plasmids. Hence a yeast expression platform has been developed at the Department of Systems Biology, DTU. The platform offers the opportunity to express genes on the chromosome in 1 to 10 copies. A comparison between the expression of CFP and RFP by the platform and by plasmids reveals the problems of plasmid expression. FACS analyses of two cell populations, expressing CFP and RFP on the separate plasmids or expressing CFP and RFP using the yeast expression platform shows expression varies greatly in a cell population based on plasmid expression compared to the yeast expression platform. When expressed on plasmids a few cells are high performers on both proteins but the largest fraction of cells is actually not expressing either of the proteins. The yeast expression platform is developed to facilitate stable expression of integrated genes. The integration sites are separated by essential genes which ensure that the integrated genes are not lost by recombination. An amplification method has been developed for the platform which enables fast integration of genes. Future perspectives involve exploring the capabilities of the platform for recombinant protein production including performance and stability studies.