gene gun transformation

BIRDD at UCRAC1.UCR.EDU BIRDD at UCRAC1.UCR.EDU
Thu Sep 29 11:50:28 EST 1994


Someone (please sign your E-mails) asked about bolistic transformation of
C. elegans. Some years ago, Phil Anderson showed that bolistic
transformation of worms was relatively easy. From memory, he used a system
(I think designed by Cetus) in which current was used to explosively boil a
drop of water, which provided the force to drive the tungsten particles. I
gather that this machine was never marketed because of legal pressure from
Du Pont. We repeated Phil's experiments using a home-made machine in which
a pulse of helium was used to provide the bolistic force. Somehow, we have
managed to lose the details of those experiments (such as particle size,
and helium pressure - which would probably be unique to our machine).
However, our approach was to collect approx 1000 washed worms on a glass
fibre filter immediately prior to shooting. Our highest transformation
rates were at pretty high kill rates (60-80 % I think). One thing we found
important was to disperse the particles immediately before shooting. We did
this by holding the bottom of the eppendorf tube containing the DNA-coated
tungsten beads directly on the probe tip of a sonicator until the
eppendoprf tube began to melt a little.
We now have access to the BioRad Biolistic gun and plan to re-try the
transformations (as a preliminary to trying to transform plant-parasitic
nematodes). We would be interested in anyones experiences, and are happy to
share ours.
David McK. BIRD
Department of Nematology and Graduate Genetics Group
University of California
Riverside, CA 92521-0415
USA

Office: (909) 787-3192
Lab: (909) 787-4586
Fax: (909) 787-3719




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