Jennifer Hicks wrote:
>> Hello!
>> I was wondering if anyone could give me information regarding the ABI 373.
> What is the difference between this machine and the 377 in read lengths?
> Are there any major differences between the two in sample preparation or in
> machine operation? How is its performance for genotyping and sequencing?
> Thanks for any insight you may be able to provide.
>> Jennifer Hicks
> Genome Center
> Brigham & Women's Hospital
> Boston, MA
I can't comment on genotyping with the 373 or 377, but we have done, and
still do a lot of sequencing on both types of machines.
We do 48 cm WTR (well to read) runs on both machines (the stretch
option) with the big dye filter wheel. We run 48 lane sharkstooth combs
on the 373 with a hacked module (we should only be able to run 36). We
used to do the same on the 377's but have upgraded to 96 lanes.
Now, for a comparison:
Sample prep is the same for both machines.
373's use thicker spacers and therefore more acrylamide and longer run
times for the same WTR length. Typically an 18 hour run on the 373 will
produce between 820 and 920 bases with an average phred Q>20 value of
>550 for M13 and a phred >20 of >500 for dsDNA. The alignable bases (those that Phrap will incorporate into an assembly) generally go all the way out to the end of the sequence (therefore 800 to 900 useable bases).
Using the same samples on a 377 you can get similar results in 9 hours
or a little less. With 48 lanes the quality is about the same as the
373, but when you switch to 96 lanes you lose some quality. At that
point you have to make a decision whether you want a little longer read
or much higher throughput.
The software for the 377 is easier to use and the new tracking software
is also much better, but then its not particularly hard to track of 48
lane gel.
We now use the 373's primarily for finishing reads on shotgun libraries
from BAC clones. We do this because the read length is longer on the 48
lane 373 than the 96 lane 377 or the 3700 and we have these resources,
so we keep them busy.
I apologize for the rambling answer. If you need a good machine for a
little bit of sequencing, the 373 is ok and probably priced right. If
you need more information, please let me know.
srlasky
--
Stephen R. Lasky, Ph.D. #
University of Washington #
Department of Molecular Biotechnology #
srlasky at u.washington.edu #
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