Taq polymerase - Taq purification (Aug/18/2008 )
I'm trying to purify Taq polymerase for the first time. This is probably a very stupid question but here goes....I'm using ammonium sulfate to precipitate the protein. Why is it that when it is spun at 13000rpm after addition of ammonium sulfate, Taq precipitates as a pellet at the bottom of the tube and also on the surface of the lysate?
Hope someone will be able to help....
If you use Ammonium precipitation the proteins are pelleted so the Taq should also be
Thanks for replying Jipes, but what I've was wondering was why all protocols recommend spinning at no higher than 13,000 rpm and this leaves some precipitate on the surface. and why is it that the protein on the surface has the highest activity? Do help if you can.
In the protocol that I have they also spin down at 12.000g for 10mn only Then they mix the supernatant with an equal volume of Tris 50mM (pH8.0) 100mM NaCl, 0.1mMEDTA, 0.5mM DTT, 1% triton containing 50% glycerol but no mention of protein clogs floating So I would just consider the supernatant