HEEEEELLLLPPPP!!-prokaryotic expression systems
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Sat Mar 31 18:00:33 EST 2001
Some of these questions you may have already addressed. Have you considered
or investigated the following?:
1. Does the plant gene have clusters of rare E. coli codons near the 5'
2. Is your protein toxic to the cells? Have you tried a strain that is
better able to express toxic proteins? Stratagene and other companies have
such cells that may help if this is the case.
3. Do you have proper spacing from you Shine Delgarno site to the gene's
ATG start codon?
4. Is the transcribed mRNA vulnerable to annealing to itself over the
ribosome binding site? (look for significant palindromic sequences flanking
the binding and start region).
5. Is your host strain able to use any inducer being used for expression?
(Example: DH5-alpha cannot have genes induced by lactose because it has
it's lac operon deleted, but IPTG can be used since it is still able to pass
through the membrane).
There are many other things that could have gone wrong. What have you done
up to this point?
ng01 at worldnet.att.net
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