what is the best miniprep

Sasha Kraev bckraev at aeolus.ethz.ch
Thu Oct 6 02:37:07 EST 1994


Dear netters, 10 days ago I posted an article (36699u$mdu...) where I tried
to introduce the observation that any commercial minicolumn is simply not
enough for a successful sequencing, that the strain is at least as important
as any of the "magic", "wizard" or other miniprep protocols. Incidentally,
I found that the Qiagen Qiaprep-spin protocol now acknowledges this fact 
as well. I have to add to this, that there are some clones ( we seem to be 
particularly blessed with those since we work with a transporter called 
Na/Ca exchanger, no matter is it from frog, dog or man.  Such clones just 
do not sequence normally with Sequenase ( cycle sequencing with
labelled primers works, however). Some of these clones have a lower than
normal copy number (and hence the yield is very low), but this does not
correlate with the background on gels. With some of these clones
we have seen a dramatic improvement in quality of sequencing, when only the
strain was changed from JM101 to XL-1 (Wizard columns were used in both cases)
At the same time I routinely sequence XL-1 minipreps without using any 
columns, and "bad behaviour" of some clones is easily traced to certain inserts,
and not only to random errors.
Appreciate any further comments on this point. 

Sasha Kraev
bckraev at aeolus.ethz.ch



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