Homologous recombination is clearly the one genetic method where zebrafish are furthest behind mice as a model system. Taking mice as a model, it would be very useful to have a pluripotent cell line that would allow transfection, selection for rare homologous recombination events, and then generation of a complete fish.
The Collodi lab has been working for a while on developing embryonic stem cells; other ideas have been to generate embryonic germ cell cultures or to use somatic cell nuclear transplantation (cloning) to allow use of fibroblasts or other existing cultures.
Another potential method could be to generate targeted double-strand breaks in an embryo (e.g. using genetically engineered zinc-finger nucleases). Jim Amatruda's, Masahako Hibi's, and Joseph Yost's lab are all trying this approach.
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We are also wokring on zinc-finger nuclease (ZFN)-mediated gene targeting, but have not yet succeeded yet. I would really like to hear from someone who succeeded in modifying endogenous and/or exogenous genes with ZFNs.
We have been working with zinc finger nucleases to stimulate HR; like Masahiko we haven't succeeded yet. I'll have a poster about this and it will be interesting to talk about possible approaches
We have tried the Drosophila I-SceI approach without concrete success so far.
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