Re: [ccp4bb] Codon Optimized Expression

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CCP4bb <-- 1999 <-- November 1999 <-- 30 November 1999

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Subject: Re: cannot read .mtz in ccp4i
From: Tim Gruene tg {- at -} SHELX {- dot -} UNI-AC {- dot -} GWDG {- dot -} DE
Date: 2008-02-02

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Subject: Re: cannot read .mtz in ccp4i
From: "Winn, MD (Martyn)" m {- dot -} d {- dot -} winn {- at -} DL {- dot -} AC {- dot -} UK
Date: 2008-02-02

Subject: Re: Codon Optimized Expression
From: Andreas_Förster docandreas {- at -} GMAIL {- dot -} COM
Date: 2008-02-02

There has been a somewhat related discussion in the lab the other day.
If some colonies might express well and other not so well, why don't I
just scoop up loads and start my culture from them? This way I'll be
more certain to get the clones that express.

The clones that don't express will dilute the yield, but I doubt they'd
outcompete the rest, would they? For me, the start-with-plenty
procedure has always worked well, but some insist on starting from
single colonies.

Andreas

James Stroud wrote:
> On Feb 2, 2008, at 2:15 AM, M T wrote:
>> One classical way to optimize expression level is to screen culture
>> conditions.
>>
>> For my proteins, I solved my expression problems by changing the
>> expression vector to a pET or changing a pET 20 to a pET 30 (if the
>> protein is toxic).
>>
>> But keep in mind that a low but folded expression is better than a
>> high expression to inclusion body.
>
> Anecdotal evidence also suggests to try picking several colonies from
> the original cloning procedure and doing test expressions with each.
> Some clones express better than others even though they should in
> principle express identically. I have no idea what the mechanism for
> this differential expression could be.
>
> --
> James Stroud
> UCLA-DOE Institute for Genomics and Proteomics
> Box 951570
> Los Angeles, CA 90095
>
> http://www.jamesstroud.com
>

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CCP4bb <-- 1999 <-- November 1999 <-- 30 November 1999

Previous message:
Subject: Re: cannot read .mtz in ccp4i
From: Tim Gruene tg {- at -} SHELX {- dot -} UNI-AC {- dot -} GWDG {- dot -} DE
Date: 2008-02-02

Next message:
Subject: Re: cannot read .mtz in ccp4i
From: "Winn, MD (Martyn)" m {- dot -} d {- dot -} winn {- at -} DL {- dot -} AC {- dot -} UK
Date: 2008-02-02