NO "WEDGE" Sequencing Gels

I have been using an alternative to wedge gels that saves acrylamide, cuts gel drying time to 20 min and gives as good or better band squashing at the bottom of the gel. It was published in BioTechniques about 3 or 4 years ago (email me if you want the reference).

The method goes as such:

Run the gel with 0.5X TBE in the top tank and normal 1 X TBE in the bottom tank. Just after the last (or in the case of one load - the only) load has run into the gel, put a half volume of 3 M sodium acetate in the bottom tank (make sure you leave enough room - I use 300 ml 1 X TBE and 150 ml 3 M Na acetate in my BRL S2 gel box).

This causes a themal gradient across the gel - the bottom is relatively cool and the top gets quite hot to touch. This causes the bands to run closer at the bottom.

A few points:

use normal acrylamide made in 1 X TBE and urea.

the gel takes half as long again to run.

if you go over 65 - 70 W, the plates crack, especially old ones.

I haven't tried not fixing it, but I see no problem.

I fix for 20 min and dry for 20 min.

I only ran 3 or 4 wedge gels but I couldn't stand the dry times, but the results appear comparable - my friends who still run 'em agree with me.

I hope this is useful.

John Nash (Internet:) Nash@biologysx.lan.nrc.ca.

Email to my other addresses is forwarded automatically,

Disclaimer: All opinions are mine, not NRC's!