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 <[EMAIL PROTECTED]>
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
