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DNA concentrations in Sequenase X'ns ??

Immune Response imnr at connectnet.com
Wed Dec 20 17:32:51 EST 1995

In article <4asort$c6b at legba.synergy.net> grant@ writes:
>From: grant@
>Subject: DNA concentrations in Sequenase X'ns ??
>Date: 15 Dec 1995 21:21:00 GMT

>Standard protocol of Sequencing reactions with unlabeled primer and labeled 
>dATP using Sequenase use a minimum of 5 microgram/reaction.
>Has anyone tried using 2 micrograms or below ??
>Will increasing the amount of primer (to say 50 pmoles/reaction) help the reaction
>produce better banding.
>If someone has an idiot proof protocol on these lines, can you please post them
>in this newsgroup.
>Thank you 

I routinely use between 0.5 and 1.0 ug of plasmid and 1 pmole of primer with 
the standard Sequenase protocol and get great sequence. 

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