Quick DNA Prep for PCR?

J. Rinehart jrineharNOSPAM at snake.eou.edu
Thu Aug 23 12:37:03 EST 2001


Hello all:

My colleagues and I have identified a PCR marker distinguishing two
species of closely-related bark beetle: Dendroctonus pseudotsugae
(Douglas Fir beetle) and D. rufipennis (spruce beetle). When we trap for
these beetles, often we get individuals in the traps that have been
damaged (usually by predators that follow the pheromone into the trap
along with their prey) to the point of being unable to identify them
morphologically, hence the usefulness of this marker. However, the
unidentifiable individuals often number in the hundreds, and so we are
looking for a rapid, non-labor intensive method of getting DNA out of
these to run our PCR reaction on.

We tried a simple crude homogenate that I never expected to work, and it
did not disappoint. We tried boiling the ground-up beetles to break up
lipid bilayers and release the genomic DNA; this also did not work.
Standard extraction methods (using Promega's Wizard genomic prep kit)
work very well, but this is way too labor and time intensive.

Sonication is not a likely option for us. Could we add a small amount of
detergent, such as Triton-X, to the crude homogenate or to the PCR
reaction? Any suggestions are welcome. Thanks.

John Rinehart
Eastern Oregon University




More information about the Methods mailing list