DNA seq Big Dyes

Harold G. Hills (hhills@iastate.edu)
Tue, 27 Jan 1998 02:08:04 -0500

We have just started to use the big dyes and are finding that we are
getting too much signal at the beginning and it fades out at the end.
Peaks beyond 500-600 bases are very small.

We are using 3ul of premix and 5 ul of the dilution buffer doing 25 cycles.

The problem does not occur with all templates, but is much more noticeable
with the big dyes then with any previous chemistry. In fact with the
dRodamine chemistry we often have too much signal at the ends of the runs.

I would appreciate any suggestions.

Hal

Harold G. Hills, Ph.D. DNA Sequencing Specialist 515 294-9585
1184 Molecular Biology Building FAX 515 294-1597
Iowa State University hhills@iastate.edu
Ames, IA 50011-3260
http://www.biotech.iastate.edu/Facilities/DSSF/ABRF For ABRF 97 DNA
sequencing tutorial.
http://www.biotech.iastate.edu/Facilities/DSSF For info about facility.