ethanol precipitation before gel purification... - (Dec/06/2006 )
hello all, i have 50 ul PCR reactions in total 12 of those tubes and i want to purify them from the agarose gel. my freind suggested that i ethanol precipitate and scale down the volume before i load on the agarose. 
what do you think?
I think direct loading without pp will work fine. Anyway, you got to purify all 12 samples, so with pp it's double task. Just get sure that you have enough amount of amplified product. I suggest purify from minigels if product is too faint.
Precipitaton before loading on gel should work, just keep in mind that concentrating your DNA might result in overloading your gel. Make sure you get rid of all EtOH though.
Another method I've done in the past is to increase DNA concentration by vacüum before loading on gel, worked fine.
Another method I've done in the past is to increase DNA concentration by vacüum before loading on gel, worked fine.
I agree.
I think it’s better to concentrate by vacuum. When you cut your bands, be sure to cut as much agarosa (with out DNA) as possible.
thank you for your replies. i dont know what you mean by vacume, we dont do it here....
about overloading the gel, maybe because it is PCR product i just have small amounts of DNA so i dont think i can overload it. i will make sure i will let all ethanol evaporate. thanx a lot again.
what do you think?How much volume reaction do you have??
When I have digestions in a large final volume, some times, I don’t concentrate or pp. I make large and thickness wells putting a little piece of tape over 2 or 3 combs and so on, and add more agarosa to have a thickness gel. This way, you’ll be able to load all your volume.
When you cut your bands, just be careful to take just your DNA and cut as much agarosa as possible.