Extension of Read Length--need new ideas

William William
Wed Jun 11 08:50:54 EST 1997


We see only a limited number of factors that influence read length on 
377's.
First, we use the 48 cm plates, and I expect this will yield some 
improvement
over shorter plates (otherwise, why make them?).  Second, the noise level 
must
be low enough that any signal attenuation will not be problematic at the 
longer reads.  Third, your plates need to be clean to avoid excessive 
smearing of the longer 'bands'.  Fourth, use the new dyes, which 
consistently
give us 600 bases with only 0-2 miscalls (exempting the first 15 bases or 
so).
Fifth, long ranger can be used, but avoid concentrations of less than 4.5 
%,
or you will have problems with band spacing (we use 4.5 to 5 %).  
Finally, I
personally recommend phenol extractions to fully deproteinize samples and
reduce potential smearing in the gel (but then I dislike columns anyway 
for 
DNA preps).  

Disclaimer:  just my $0.02--your mileage may vary.

Bill Buikema

William J. Buikema, Ph.D.   Phone: (773)702-1081   e-mail: 
hetman at uchicago.edu
Dept. Molecular Genetics & Cell Biology          /  / --- --- /|/|  /|  
/| /
Univ. of Chicago, 920 E. 58th St.               /--/ /-   /  / / | /-| / 
|/
Chicago, IL 60637 USA       Fax: (773)702-3172 /  / /__  /  /    |/  |/  /
http://cancer-seqbase.uchicago.edu/

-- 
William J. Buikema        Phone: (773)702-1081   e-mail: 
hetman at uchicago.edu
Dept. Mol. Genet. & Cell Biol.                   /  / --- --- /|/|  /|  
/| /
Univ. of Chicago, 920 E. 58th St.               /--/ /-   /  / / | /-| / 
|/
Chicago, IL 60637 USA       Fax: (773)702-3172 /  / /__  /  /    |/  |/  /



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