E.coli rare codons

ijiwaru at wheel.dcn.davis.ca.us ijiwaru at wheel.dcn.davis.ca.us
Tue Sep 3 23:22:09 EST 1996


In article <322E1166.3687 at jcu.edu.au>, Lynn Hughes
<lynn.hughes at jcu.edu.au> wrote:

> Hi all,
> I am attempting to clone my DNA fragment into an E.coli expression 
> system.  My DNA contains quite a high percentage of 'E.coli rare codons'. 
> Will this lead to low expression levels of my protein, if so any ideas 
> how I can get around this problem, for example any special feature I 
> should look for in the expression system I choose.
> 
> Thanks for your help
> 
> Lynn Hughes

Lynn,

it may not be too much of a problem, depending on your protein.  You may
want to try a couple different things.  One, put your expression system
into context of a low copy number vector and count on a long, slow
expression.  In this way, you don't tax the system by asking for a whole
bunch of the rare tRNAs all at once.  Two, a variant of one, when you do
your expression, instead of a high level of induction for a short period
of time, lower levels of inducer for a long period of time.  This will
work quite well if your protein is reasonably stable in coli, if its not
toxic, and if you're patient.  Unless the particular vector system has the
rare tRNAs in it or you're going to change the unfriendly codons, I don't
think there's too much else you can do but take the lower levels of
expression and try to get workable quantities of protein.

Good luck,

Lyle Najita
Plant Pathology
University of California - Davis



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