ABI sequencer: reaction clean-up ??

Jason E Stewart jasons at bmc.uu.se
Fri Jan 22 17:23:27 EST 1993


In article <1993Jan22.122927.16338 at news.unige.ch>, mike at medsun.unige.ch
(michel morris) wrote:
> 
> 	We are about to do our first sequencing on an ABI 
> sequencer, using their DyeDeoxy/Taq pol. protocol. 
> 	ABI recommend (insist!) on cleaning up the reactions 
> before loading, preferably with their own spun columns - surprise, 
> surprise.
> 	Is this really necessary? How do the other protocols
> they mention, such as phenol/CHCl3 or isopropanol ppt. compare?
> What is the quickest way to get the reactions onto the machine 
> (while still getting sequence off...)?
> 
> tia,
> Mike Morris

Absolutely critical to get rid of the free nucleotides. Although we have
only used it a few times I would recommend the double phenol extraction and
warn against the iso-prop. precip. I would also recommend climbing on your
suppliers heiny and get him to install the T7 terminator kit as soon as it
comes out. With the fluorescein terminators, no cleanup is necessary.

Good luck,

Jason

----------------------------------------------------------------
Jason Stewart    	                      Father at large
Department of Medical Genetics
jasons at bmc.uu.se



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