Long PCR
AShiflett
ashiflett at mbl.edu
Thu Mar 25 16:13:37 EST 2004
Hi! I am attempting to amplify an approximately 12 kb piece of DNA
that is near the telomere of a chromosome. I have used both Herculase
from Promega and the Elongase enzyme from Invitrogen. In both cases,
I do not obtain a product of the expected size. I followed the mfg
directions, and performed optimization for primers, annealing temp,
and Mg concentration. I use approximately 100 ng of genomic DNA which
is not sheared and looks great on a gel. Does anyone have any tips
for amplifying long pieces or DNA, or can recommend a good polymerase
that has been successful for difficult PCRs? Any advice is welcome
and appreciated.
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