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