colin at pombe.usask.ca
Thu Aug 12 13:37:34 EST 1999
Joan Marie Shields wrote:
> XL wrote:
> >> I am running a lot of DNA sequencing gels in my lab. One of the reagents we
> >> use routinely is the TBE. We usually make a 10X stock and make dilution from
> >> there. As many of you may already notice that 10XTBE will precipitate after
> >> the stock is used/opened. I am trying to get some chemistry explanations
> >> about this precipitation. Is it because of the solubility, pH or chemical
> >> reactions with air? Thank you.
> Colin Rasmussen <colin at pombe.usask.ca> wrote:
> >I think the borate has a tendency to ppt. with time. We found in the
> >past that autoclaving it helps for some reason. Alternatively, make 5 x
> >TBE. That always solves the problem. We typically run our gels in 0.5
> >x TBE so you won't wind up making any more than you are now.
> I was having trouble with it precipitating as well - autoclaving seems to
> have solved that problem.
> I will say regarding the 0.5x - I typically run fairly high percentage
> agarose gels (2-4%) and I've found that if I use 0.5x TBE that my bands
> aren't nearly as sharp. I've gone back to using 1x TBE and have had much
> better results.
What voltage do you run at. We've found that at 100v on a minigel,
sharpness is fine, although I must admit we don't run that high
percentage. When I need to separate smaller things I usually switch to acrylamide.
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