Cycle Sequencing of cosmid templates?
Tue Jun 24 08:17:47 EST 1997
Does anybody know of a reliable protocol for sequencing cosmids by cycle
sequencing? Of special interest for me are the amounts of DNA template
and primer oligonucleotid to be used in the cycling reaction.
I tried the sequencing reaction with standard template amounts for
plasmid DNA (about 500 ng which yielded good results in our standard
plasmid sequencing reactions) and with significant higher template
concentration (about 2 ug) but got no usable sequence using standard
sequencing protocols referring to the ABI 373/PE ABI Prism Cycle
In addition, I am also looking for a cosmid DNA purification method
which yields good amounts of (sequencing-grade) clean DNA. Up to now, I
used QIAGEN column purification but this method resulted in very low
Every help is greatly appreciated.
Thank you in advance,
Institute of Human Genetics
University of Saarland
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