Switch to the use of taurine buffer in the gel and as running buffer.
solution to the problem has been given to Applied Biosystems>. 20X
buffer: 1.78 M Tris, 0.57 M Taurine, 0.01M EDTA. To make 2 liters we use
432 g Tris Base, 144 g Taurine and 8.0 g DiSodium EDTA (FW 372.24).
Taurine buffer can be purchased as a 20x solution from Amersham Catalog #
US 75827. Taurine buffer does not precipitate, goes into solution easily
and we never bother to check the pH. Our Instrument Sales rep gave
supplies to Laura Livingstone at UNC-Chapel Hill for solving this problem.
In article <5cnrkv$icn at net.bio.net>, Dr Tim Sawbridge,
>T.Sawbridge at forbio.com.au wrote:
>>> Dear Sequencers,
>> we have been seeing stretching of peaks from around 190
>> to 210 bp across the gel on a 377 with resulting loss of resolution. The
>> peaks return to their normal width after this so it seems to be a
>> localised problem at around these fragment sizes. We are doing 3.5 hour
>> runs on 36cm plates using dye-terminator chemistry with the ABI kit on
>> double stranded plasmid DNA. Has anyone seen this and know what might be
>> causing it?
>> Thanks in advance
>> Tim Sawbridge
>>>>T.Sawbridge at forbio.com.au>>>We have been having the exact same problem (377, DT, 36cm, 4X run). We
>thought it was a problem in the Gel reagents but it continued after
>changing them. We acid cleaned the plates but that didn't help.It happens
>on both a 4% polyacrilimide gel and on the Long Ranger gels. We usually
>pour our gels in the evening and run them the next morning. Maybe these
>clues will help. Please respond with some help if you figure this out.
>>Thanks, Charlie Demma Demma at welchlink.welch.jhu.edu
Harold G. Hills, Ph.D. DNA Sequencing Specialist 515 294-9585
1184 Molecular Biology Building FAX 515 294-1597
Iowa State University
Ames, IA 50011-3260 hhills at iastate.eduhttp://biotech.zool.iastate.edu/Biotech/Facilities/DSSF/homepage.html