Re: [ccp4bb] insect expression system

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

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Subject: Re: insect expression system
From: "A {- dot -} Radu Aricescu" radu {- at -} STRUBI {- dot -} OX {- dot -} AC {- dot -} UK
Date: 2008-05-28

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Subject: Re: insect expression system
From: Vangelis Christodoulou e {- dot -} christodoulou {- at -} NKI {- dot -} NL
Date: 2008-05-28

Hi Yongfu,

I couldn't agree more with Radu. We had great success in expressing mg
amounts of a secreted protein in HEK293 cells (with Radu's help :-) .
The same protein was initially expressed in Sf9 cells but with much
lower yields.
Furthermore, we could very easily generate a stable HEK293 cell line
expressing the same protein (at similar levels with transient
transfections) with the Flp-In system in just a couple of weeks. We
also have a LIC vector which is compatible with the Flp-In system.

Hope this helps,
Vangelis

--
Vangelis Christodoulou
Analist C
Anastassis (Tassos) Perrakis lab
The Netherlands Cancer Institute (B8.038)
Plesmanlaan 121, 1066 CX, Amsterdam
The Netherlands

tel: +31 (0)20 512 1963
Website: xtal.nki.nl/Tassos_group/

On May 28, 2008, at 7:36 PM, A. Radu Aricescu wrote:

> Dear Yong-fu,
>
> Why would you worry about insect expression systems if you already
> can secrete your constructs in mammalian cells? For such proteins,
> transient expression in HEK cells for example gives higher yields
> than baculo, is faster, cheaper, you can nicely control
> glycosylation, easily do Se-Met labelling and so on.
>
> Here are some references (PMID): 17355862, 17001101, 16823037,
> 11788735, 16082028.
>
> Radu
>
> ------------------------------------------
> A. Radu Aricescu, PhD
> University of Oxford
> Wellcome Trust Centre for Human Genetics
> Division of Structural Biology
> Roosevelt Drive, Oxford OX3 7BN
> United Kingdom
> Phone: +44-1865-287551
> Fax: +44-1865-287547
>
>
> ---- Original message ----
>> Date: Wed, 28 May 2008 10:40:16 -0400
>> From: Yong-Fu Li
>> Subject: [ccp4bb] insect expression system
>> To: CCP4BB@JISCMAIL.AC.UK
>>
>> Hi,
>>
>> I searched my mail box for possible answers you have
>> given but couldn't find any. The question is about
>> selecting insect expression systems for mammalian or
>> viral glycosylated proteins. There is confirmed
>> expression of the target proteins in mammalian
>> cells. They are secreted into the media.
>>
>> 1) Baculovirus system
>> 2) Drosophila system
>>
>> Which system would you recommend for high expression
>> and convenience?
>> Has anyone ever compared those systems side by side?
>>
>> Any suggestions or references are greatly
>> appreciated.
>>
>> Yongfu Li

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

Previous message:
Subject: Re: insect expression system
From: "A {- dot -} Radu Aricescu" radu {- at -} STRUBI {- dot -} OX {- dot -} AC {- dot -} UK
Date: 2008-05-28

Next message:
Subject: Re: odd waters
From: Bernhard Rupp bernhardrupp {- at -} SBCGLOBAL {- dot -} NET
Date: 2008-05-28