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I will be traveling to University College London (England) to work for
two years starting in May 1995. My research involves the generation of
an unusual variety of 'gene knockout' mice. Traditional researchers in
this area use pronuclear microinjection of single cell embryos (usually
microinjection under visual guidance for each and every embryo). Among
other possible techniques I am considering the use of electroporation of
embryonic stem cells and FACS screening to select low frequency (perhaps
1 in 1000) cells exhibiting the appropriate DNA excision or integration
events. Do you think it is possible to use FACS to detect an appropriate
homologous recombination event with great specificity (isolating the few
cells which hybridize [or fail to hybridize] to a synthetic DNA probe) --
yet maintain the viability of these embryonic stem cells? Would it be
possible to provide any references to recent literature (or people who
are doing this kind of work) where viable embryonic stem cells are being
sorted based on a synthetic DNA probe -- and yet maintain viability?
I read your 2nd edition book some time ago. I just purchased a copy of
your 3rd edition book. I like the organization of the new book. But,
I rather miss the practical here is how to build one yourself approach
of the previous edition. I understand most readers didn't bother to try
to build an instrument for their own use. But, I built many Scientific
American Amateur Scientist projects when I was a child (far too long ago)
-- I rather liked seeing some of the same kind of advice (certainly at a
far more professional level) in your previous edition.
In any case, thank you for your kind help and/or advice regarding the
possibility of using FACS to screen viable embryonic stem cells for an
appropriate engineered gene sequence modification event.
Thanks, AJ Annala
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Alexander-James Annala
Principal Investigator
Neuroscience Image Analysis Network
HEDCO Neuroscience Building, Fifth Floor
University of Southern California
University Park
Los Angeles, CA 90089-2520
Telephone: (213)740-3406
FAX: (213)740-5687
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