PCR Amplifying 2-5Kb fragments
me at privacy.net
Fri Jun 25 03:20:06 EST 2004
Historians believe that in newspost <10dkcsg6kjqrcb9 at corp.supernews.com>
on Wed, 23 Jun 2004, Phil McClean <mcclean at cableone.net> penned the
following literary masterpiece:
> Does anyone have a procedure in which they can routinely amplify via PCR
>fragment 2 to 5 kb in length.
Do it all the time from bacterial genomic.
>What are some of the critical factors that
>you have encountered.
It really depends upon the complexity of the genomic source. Human
xsomal is much more difficult than a bacterial PCR. GC rich is much
harder than a 50% GC etc.
More enzyme can help i.e. doubling or even quadrupling at times. 4-5kb
on human genomic is pushing it with standard Taq. Putting in say 2.5u
Taq with 0.125u Pfu and using the Pfu PCR buffer can make a huge
I love deadlines. I especially like the whooshing noise they make as
they go flying by.
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