5 replies to this topic

[saraarasus]

HI,
HAS ANY ONE TRIED TO EXPRESS PGEX in the eucayotic cells or any recomendation?

[pras45]

saraarasus, on Jan 27 2009, 02:49 AM, said:

HI,
HAS ANY ONE TRIED TO EXPRESS PGEX in the eucayotic cells or any recomendation?

Yes, what is your specific question

[rkay447]

The pGex vectors are not intended for expression in eukaryotic cells and does not contain an appropriate promoter.  These vectors are designed for expression of your protein in bacteria for purification.

[pras45]

saraarasus, on Jan 27 2009, 01:49 AM, said:

HI,
HAS ANY ONE TRIED TO EXPRESS PGEX in the eucayotic cells or any recomendation?

Look up if your protein has rare codon (if mammalian), if so then Novagen's rosetta BL21 strain would be a good option. I have used BL21(DE3)pLysS (auto lysozyme production) and has given me fairly good result.

[kasturi]

elution of basic proteins
I am using the PGEX system to purify a basic protein of PI 9. I am using a denaturing 2M urea based protocol that i came across in a paper. The protein seems to be binding the column but when I try to elute it with PBS having 10mM glutathione + 2M urea nothing comes off. Can someone suggest some variations I could try to recover it from the column. Will changing PH conditions help?

[swanny]

pras45, on Feb 5 2009, 03:42 AM, said:

saraarasus, on Jan 27 2009, 01:49 AM, said:

HI,
HAS ANY ONE TRIED TO EXPRESS PGEX in the eucayotic cells or any recomendation?

Look up if your protein has rare codon (if mammalian), if so then Novagen's rosetta BL21 strain would be a good option. I have used BL21(DE3)pLysS (auto lysozyme production) and has given me fairly good result.

I think you'll find that Rosettas are prokaryotic...

So the basic answer to the original question is in posting from rkay.