Hi,
you did not mention if you just look for soluble expression.
However you could try an other fusion construct like MBP or GST-Tag to check
if the RNA do some strange things (secondary structures).
There are more strains available. For some unknown reason some strains work
some not. Sometimes ther is also a difference when you use an other media TB
instead of LB.
Maybe you know someone who has experience in Bacillus Expression systems.
Nothing is better than the own host.
Might also be possible that your protein is toxic for E.coli.
Christian
Am Mittwoch 14 April 2010 21:30:05 schrieb Amit Kumar:
> Hi all,
>
> Sorry for a non-ccp4 query.
>
> I will highly appreciate your comments on the expression:
>
> I am trying to express HIS tagged ~60 kDa cytosolic protein from Bacillus
> in E.coli pET expression system. I tried codon+ strain, time of
> expression and various temperatures ( 18, 25, 30, 37C) for the
> expression. Neither full-length nor truncated form of the protein was
> found to be expressing (also checked by western blot). What other
> parameters should I explore to get the expression in Escherichia coli?
>
> Thank you very much for your comments.
>
> Amit K.
>
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