Thursday, October 16, 2008

SB buffer for DGGE?

I recently received this question:
Hi Stefan,
I am new to this blog and stumbled upon it. Have you used the SB buffer in the DGGE apparatus? If so what have your run times and concentrations been? I have been using SB buffer with standard agarose gels but I am still using TAE buffer in our DGGE set up at 150V for 10h. Just curious and interested in saving some time.

-- My response:
No, I haven't tried it. I am now in a lab without a DGGE, so I haven't had a chance. Theoretically, it should work and it should significantly reduce the cost of making buffer. I doubt it will make a big difference in the running time, because the limitation on voltage in a DGGE doesn't have much to do with the gel heating up (as in agarose gel electrophoresis).

Some thoughts - make sure to use SB buffer when making the acrylamide stock solutions. If you (or anyone else) tries SB buffer in place of TAE, please email me about the results and I'll post it.


P.s. I have received the following comment from Michael (see below). His response would indicate that SB buffer is NOT appropriate for DGGE. C'est la vie.

"In fact I also have quite bad experience with SB buffer. Apart from the worse separation, it seems that SB somehow modifies gel's structure, making it softer and more fragile. Moreover, gels submerged into the staining solution significantly increase their dimensions, which suggests some kind of inhibition of the poliacrylamide polimerisation."