QIAquick Gel Extration Kit - (Sep/08/2005 )
What does everyone get for % recovery when they use the kit with microcentrifuge? We spin at 12,000 instead of 13,000, could that be why we get 20% recovery?
No, using the quiagen system you should be able to reach a recovery of at least 80%, no mather whether you spin at 12000 rpm or 13ooo rpm in a table centrifuge.
What size DNA are you trying to purify? Also, what buffer are you using to elute. This system is pH dependent, so if you are using H20, you need to bump up the pH to get efficient elution.
Yes, what size is the product? I've had decent recovery with QIAquick but I prefer Zymoclean, you elute to a smaller volume which I've found is better for cases when I've had low recovery with QIAquick.
For large fragments, it might help to pre-heat (50-60 degrees celcius) the elution buffer.
My experience is that when extracting DNA from gels, I also loose a lot of my DNA, never really bothered to calculate how much.
It will help to increase the % recovery if you run your elute through the filter again. Usually not all the DNA get eluted during the first elution so the second time will help. It seems to work well for me and I always elute the DNA twice.
It will also help to heat up the Buffer EB or Tris.Cl or pH 8.0 H2O to 80C if your DNA is large.
Is the ethanol that you added to the wash buffer 200 proof or denatured? Only use 200 proof ethanol or your yields will be much lower.
I lose all PCR product by using 12000 rpm , so I try to reduce the speed to 8000 rpm. It's ok