DNA recovery by phenol/chloroform extraction or GeneClean

Wilson netson at uxmail.ust.hk
Tue Feb 11 07:46:38 EST 1997



On Mon, 10 Feb 1997, Sai wrote:

> that's a question i would like to know the answer to as well...i've
> consistently been getting low yields of recovery after the gene clean
> process (even if my gel fragment is pretty concentrated).  does anybody
> here have a speed vaccuum step at the end to get rid of all the NEW wash? 

It may not necessary use a speed vaccuum to remove NEW wash, just use
pipette to remove the NEW wash.  I have heard that too dry of the pellet
may make the elution difficult.

> because i suspect that might be where i may be losing some of the bound
> DNA (although i cant see why; DNA shouldnt be volatile)...anywayz, any
> other suggestions to get purified fragment would be much
> appreciated...thanks...

The step which vortex the pellet after adding the NEW wash may decrease
the yield of recovery. (I guess).  Therefore, I do not vortex the pellet,
just incubate the pellet at room temp. for 5 minutes.  After eluting the
DNA to TE, I do ethanol precipitation to remove remaining NaI.
By the way, I have found a problem at the last step of elution of DNA from
glassmilk.  I often remove the eluted DNA with little amount of glassmilk
even after centrifugation of the pellet.  How can I prevent it?

Wilson 




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