A COPY-NUMBER-CONTROLLED EXPRESSION VECTOR FOR THE FISSION YEAST SCHIZOSACCHAROMYCES-POMBE

A novel, expression vector for the fission yeast Schizosaccharomyces p ombe carries the neomycin-resistance-encoding gene regulated by the SV 40 early promoter, and its copy number is controlled by the level of G eneticin (G418). Foreign gene expression is driven by the human cytome galovirus (hCMV) promoter which is transcriptionally active in S. pomb e. Moreover, the vector expresses foreign genes at high levels, due to the 5'-untranslated region (5'-UTR) containing an A + T-rich sequence of about 50 nucleotides located between the TATA box of the hCMV prom oter and the start codon. Recombinant human lipocortin I was produced at levels of up to 50% of the total soluble protein in the presence of 100-200 mu g/ml of G418 in the media. Southern and Northen blotting s howed that this high level of expression was due to an increase in cop y number induced by G418, the high transcriptional activity of the hCM V promoter and the high translational efficiency of the 5'-UTR. We mod ified the vector into an 'ATG vector', named pTL2M, that maintains the 5'-UTR optimized for gene expression and into which any foreign gene, whose exact sequence is known, can be easily inserted.