Glowing Blue DNA Gels

Alistair Forrest forrest at biosci.uq.oz.au
Sun Apr 17 18:42:39 EST 1994


skralyf at cae.wisc.edu (Frank Anthony Skraly) writes:

>What the . . . ?

>The last 2 times I ran agarose gels (TAE buffer, stained in dilute ethidium
>bromide/water), I put them on the UV box and they glowed 

>	BRIGHT BLUE ALL OVER.

>This made it very difficult to see the orange EtBr/DNA bands. 

>The glowing would recede to bright blue spots on the gel after long destaining
>periods in water, but would never go away.

>Today I ran a well-behaved gel, but it had one blue dot.

>Sometimes we use 1X EtBr solutions that have been sitting around for a day or 
>two (in the dark), and I wonder if the EtBr turns into something else that 
>fluoresces BLUE.  We've always done this, though, and we've never seen the BLUE
>effect.

Just sounds like you've got too much EtBr in your running buffer. To overcome
this you could try including the EtBr in your gel (we use 20ul of 10mg/ml in
a 40ml gel) or you could also try prestaining the gel in a concentrated solutionof EtBr before running. The advantage of this system is that as the EtBr is alsoelectrophoretically mobile and smaller than the DNA, it will run off the end of the gel. The EtBr is then diluted in the running buffer and as such it won't
matter if the Et Br is recirculated to the front of the gel as by now it is
so dilute it shouldn't pose any problem.

Alistair Forrest




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