Cloning yeast ORF: any sequences upstream of ATG needed?

05 May 2013 1 849 Report

I am cloning a yeast ORF downstream of the ADH1 promoter. For all of our MAMMALIAN work we ensure there is a Kozak sequence upstream of the ATG. But what about yeast? Will I get good expression if I simply clone from the start codon, or is a small amount of upstream sequence (or a specific sequence) necessary for good expression of the protein?

Chung-Yun Chao

I would include an yeast Kozak sequence upstream. Some literature describes a significant difference in expression, although we have had instances where we did not include a Kozak sequence and expression was still observed.

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