Polyethylenamine and Ni-NTA/anion exchange

Dr. Duncan Clark d_clark at nospam.demon.co.uk
Thu Mar 15 05:36:57 EST 2001


In article <Jonathan_Kurtis-1403011125400001 at 10.137.37.205>, the eminent
Jonathan Kurtis, MD/PhD at International Health Institute wrote
>As part of a purification protocol, I have been precipitating nucleic
>acids from recombinant E.coli lysates with polyethylenamine. (Add
>polyethylenamine to 0.5%, incubate on ice for 20 min, spin 20,000 x g for
>20 min and collect sup).

The problem with PEI is that unless you have some salt present, >0.1M
NaCl, then you stand a very good chance of ppting the protein you are
looking for. The best thing to do is to titrate the PEI and monitor by
PAGE. If your protein is a DNA binding protein then it may also come
down even in the presence of some salt.

Most procedures with PEI also require that you remove the PEI before
loading a chromatography column (not always the case but a general
rule). It cannot be dialysed out so the standard method is AmmSO4
pptation of the protein leaving PEI in solution. 

Duncan


-- 
The problem with being on the cutting edge is that you occasionally get 
sliced from time to time....

Duncan Clark
GeneSys Ltd.
Tel: +44(0)1252376288
FAX: +44(0)8701640382
http://www.dnamp.com
http://www.genesys.demon.co.uk




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