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Development

Development. What causes cell differentiation?. Molecular Markers. Homeobox Genes. Transcription factors encoding the DNA binding homeodomain. Expressed in temporal and spatial specific patterns during development. Cell differentiation is disrupted when they are mutated. Adult Spinal Cord.

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Development

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  1. Development What causes cell differentiation?

  2. Molecular Markers

  3. Homeobox Genes • Transcription factors encoding the DNA binding homeodomain. • Expressed in temporal and spatial specific patterns during development. • Cell differentiation is disrupted when they are mutated.

  4. Adult Spinal Cord

  5. Neural Tube Development How are the expression patterns for homeodomain proteins created?

  6. Lbx1 Expression Pattern Pax3•Lbx1

  7. Project Goal Identify the tissue specific enhancer that creates the Lbx1 expression pattern in the dorsal horn of the spinal cord.

  8. Mouse Lbx1 Locus

  9. 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.

  10. In Ovo Electroporation: GFP ß-actin Enhancer GFP

  11. Instruments

  12. Positive Control: ß-Actin GFP Expression Vector

  13. Asymmetric Expression of GFP

  14. In Ovo Electroporation: LacZ Cytomeglavirus Enhancer LacZ

  15. Positive Control: ß-Galactosidase Reporter Gene

  16. Results

  17. Conclusion • Tissue specific enhancer does not exist within our search area. • Can now start searching upstream and downstream.

  18. Acknowledgements Dr. Michael Gross Dr. Kevin Ahern Howard Hughes Medical Institute

  19. Questions?

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