I am trying to express a bacterial protein in Saccharomyces cerevisiae. The protein is expressing well in BL21DE-3(E.coli). It is not getting expressed in S.cerevisiae. Is it worth going for codon optimization and gene synthesis for expression of that protein in S.cerevisiae.
You need to check the codon usage of your bacterial gene and compare it to that of E. coli and S. cerevisiae. Based on the results you may give it a trial or not. If you opt for codon optimization I would optimize it for expression in both organisms, S.cerevisiae and E. coli.
Hi there,
Is it a soluble or membrane protein? What kind of vector do you use? What kind of Sc strain do you use? As already mentioned, codon optimization may considered as a first step if codon use is very different in your bacterial ORF.
Dear liger and stolz,
thanks for the inputs. I am using YSH (a leu2-3/112 ura3-52 trp1-92) S.cerevisiae strain for the expression. I am trying with commercial BY4741 strain as well. It is a soluble protein (an enzyme) involved in nitrogen metabolism. I am using both integration (pRG235) and episomal vector (p426GPD). I have analysed the gene and it contains rare codons and hence the difference in codon usage.Does optimizing the gene for S.cerevisiae helps for expression.
Are you sure it is not expressed? Do you follow it with an enzymatic assay or just on the gel?
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