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| Transgenic Transposon GFP expressing tadpole. |
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SCREENS
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Sleeping Beauty Mutagenesis
My laboratory has focused on the use of transposons for insertional mutagenesis screens. We are using the Sleeping Beauty transposase to catalyze the insertion of synthetic transposons into the frog genome. We have developed transposons that function as enhancer traps (minimal promoter constructs driving expression of GFP) and as gene traps (promoterless GFP reporter with a 5’ splice acceptor site).
We are using two approaches to generate enhancer and gene trap animals; (1) co-injection of the transposon DNA with mRNA encoding the transposase and (2) transposon hopping. Transposon hopping relies on the “cut and paste” activity of Sleeping Beauty to remobilize an existing transposon to a new site in the genome. Male frogs that carry a transposon and also harbor a transgene encoding Sleeping Beauty expressed from a testis-specific promoter are out-crossed to wild type females. Sleeping Beauty is expressed in the spermatocytes of the double transgenic males and catalyzes the excision and reintegration of the target transposon to a new site in the genome. Tadpoles will be scored at various stages of development for novel fluorescent protein expression patterns.
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Transposon Gene Traps
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Transposon Enhancer trap
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Retroviral Insertions
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Transgenic Lines
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