Plus check the insoluble part or the whole cell.
Date: Sat, 5 May 2012 16:30:06 -0400
From: jubo...@jhsph.edu
Subject: Re: [ccp4bb] no expression
To: CCP4BB@JISCMAIL.AC.UK
Some very stupid questions from my side:- it is an expression vector and you
are in frame with your gene of interest ?- you are using E. coli strains
suitable for expression ?- have you tried adding glucose to the media to avoid
leaky expression in case your protein is toxic to E. coli ?- the wild type
expresses well ? In the same vector & strain ?- how do you determine if it was
expressed ? Western blot of tag ?
Jürgen
Sent from my iPad
On May 5, 2012, at 16:17, "Jahan Alikhajeh" <ja...@graduate.org> wrote:
Dear friends,
I am sorry for off-topic though it may be related indirectly!
I mutated a 60kDa protein (changing from X-->Pro). After doing all the steps, I
sequenced the expression vector. It seems everything is fine, even with
promoter, but, I can't express it. I tried different Tempratures, IPTG (e
with new Stock), media, but they didnot make any difference. Any suggestion is
highly appreciated.
Best,
Jahan
