I tried to express protein from glycerol stock (pGEX vector and BL21 strain) that is approximately 10 years ago. There is no expression in optimum conditions. What reasons could exist for this? And what is the solution?
Christina R Bourne
Try purifying your plasmid, verify it is still OK (agarose gel, restriction digest, even re-sequence), and transform to fresh BL21 cells.
There are lots of reasons - the cells are no longer viable, they have repressed the plasmid, mutations have been introduced, general lab gnomes / unlucky, etc.
Good luck!
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