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Development What causes cell differentiation?
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Molecular Markers
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Homeobox Genes 1.Transcription factors encoding the DNA binding homeodomain. 2.Expressed in temporal and spatial specific patterns during development. 3.Cell differentiation is disrupted when they are mutated.
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Adult Spinal Cord
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Neural Tube Development How are the expression patterns for homeodomain proteins created?
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Lbx1 Expression Pattern Pax3Lbx1
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Project Goal Identify the tissue specific enhancer that creates the Lbx1 expression pattern in the dorsal horn of the spinal cord.
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Mouse Lbx1 Locus
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Delivery of Reporter Constructs to Embryonic Cells Different experimental models can be used. –Mouse common but difficult, expensive. –Chick easy, inexpensive. Our method of choice: electroporation of chick embryos. Method is relatively new.
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In Ovo Electroporation: GFP ß-actin Enhancer GFP
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Instruments
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Positive Control: ß -Actin GFP Expression Vector
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Asymmetric Expression of GFP
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In Ovo Electroporation: LacZ Cytomeglavirus Enhancer LacZ
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Positive Control: ß-Galactosidase Reporter Gene
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Results
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Conclusion Tissue specific enhancer does not exist within our search area. Can now start searching upstream and downstream.
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Acknowledgements Dr. Michael Gross Dr. Kevin Ahern Howard Hughes Medical Institute
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Questions?
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