The best rule that I found is 5-10ng per 100bp,except for PCR products with
high GC content or secondary structures where I use twice this quantity.
This rule give consistent results with well purified PCR products.
Jean R.
Dear Autosequencers,
>>We are in a bit of a dilemma - Does anyone have a good "rule of thumb"
>for the amounts of PCR product required for sequencing, per 100bp of
>product length? We are using Perkin Elmer Taq FS labelled terminator
>chemistry. We have found a lot of the manuals were written for
>ordinary Amplitaq. The PE kit instruction manual suggests different
>amounts from their Comparative PCR sequencing booklet for the same
>enzyme. Our users that are getting good direct sequencing results are
>not confident of the accuracy of their estimations. Confusing, hey?
>--
>______________________________________________________________________
>>Jennifer Cassady
>Manager
>DNA Sequence and Analysis Facility
>The University of Queensland
>St. Lucia, Qld. 4072
>AUSTRALIA.
>>>Ph: +61-7 3365 4601
>Fax: +61-7 3365 4388
>email : cassady at biosci.uq.oz.au
Jean Renaud
Recherche en Sciences de la Vie et de la Sant=E9
Pav. Charles-Eugene Marchand
Universit=E9 Laval
Qu=E9bec,(Qu=E9.),Canada
G1K 7P4
t=E9l.:418-656-2552
fax.:418-656-7176
Internet:jrenaud at rsvs.ulaval.ca
