Sunday, January 13, 2008

Alternative Electrophoresis Buffer to Replace TAE or TBE

- IMMEDIATE LINK TO GUIDE -

I am revising this submission to be a little clearer. I would like to advocate the use of three new electrophoresis buffers. Links to the relevant manuscripts are below. Also found below is a link to the company “Faster-better-media” selling these buffers.

(1) SB Buffer – 10 mM sodium boric acid (1X working solution is either 5 mM disodium borate decahydrate or 10 mM NaOH adjusted to pH 8.5 with boric acid; up to 50X concentrates can be made. 20X concentrate should have a pH of 8.0 → for more details see the manuscript below)

(2) LA Buffer – 5 mM lithium acetate

(3) LB Buffer – 1 mM lithium boric acid (lithium hydroxide pH buffered to pH ~8.5 with boric acid)

The advantages of these buffers are multiple; (a) they generate much less heat than TAE or TBE buffers, (b) they can be run at much high voltages (>300V in some cases) without fear of gels melting, (c) they can be much cheaper than TAE or TBE, (d) they do not interfere with standard applications such as band excision, and (e) they last longer before needing to be replaced.

The authors recommend that for standard electrophoresis applications, buffer SB be used. We are running gels with this buffer at 300V. For longer DNA fragments (>3 kb), buffer LA should be used. I have not yet tried this buffer. LA is also recommend for replacing MOPS in RNA separations. Buffer LB can be run at even higher voltages than SB, and is appropriate for separation of small DNA fragments.

Faster-Better-Media website


Brody JR, Kern SE. Sodium boric acid: A tris-less, cooler conductive medium for DNA electrophoresis. BioTechniques 2004; 36:214-216

Brody JR, Kern SE. History and principles of conductive media for standard DNA electrophoresis. Analytical Biochemistry 2004; 333:1-13.

Brody JR, Kern SE. Ultra-fast high-resolution agarose-electrophoresis of DNA and RNA using low-molarity conductive media. BioTechniques 2004; 37:598-602.

Cheers,
Stefan

101 Comments:

At 9:48 AM, Blogger Dustin said...

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.

Dusty

 
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At 8:05 PM, Anonymous Anonymous said...

we have tried SB buffer and run it at 200 V but our 1KB DNA ladder did not resolve properly. i.e high mol wt bands were clumped in top only. how can we solve this

 
At 8:16 PM, Anonymous Panic Attack said...

The revision did provide a clearer picture. The buffer issue is a challenge, but it needs to be resolved. Will await the update of this matter.

 
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