I used two expression plasmids, pESC-His and pESC-Trp, to express 4 genes in Saccharomyces cerevisiae. Initially when I expressed these genes, I found that the strain was producing the compound of interest, a sesquiterpene-alcohol. About one year later I had to repeat this experiment, so I revived the strain from cryo stock and repeated the fermentation as before. Surprisingly no compound was produced. I checked all the chemicals and media components used and didn't find a problem. Finally, I retransformed the yeast with the expression vectors to obtain fresh strains. More surprisingly, the compound is still not formed in these fresh strains. I would really appreciate anyone who can help me with this problem. Thank you
I find hard to believe that all the cells in the stock would stop working. It is easier to explain it with a mistake in labeling the stocks.
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