This proposal describes a 5-year training program for the development of the principal investigator into an independent investigator in the field of comparative medical genetics. James Kehler received his VMD in 2002, and is working on his Ph.D. thesis in Cell and Molecular Biology at the University of Pennsylvania. During the first 3 years of the proposal, the principal investigator will receive superb training in molecular and stem cell biology in the laboratory of Hans Scholer, his thesis advisor and sponsor of this proposal, at the Center For Animal Transgenesis and Germ Cell Research. He will have the opportunity to learn new transgenic techniques in mice and large animal species from additional researchers at this unique center. During the last 2 years of the training grant, the principal investigator will work in the laboratory of Dr. Stephen O'Brien, co-sponsor of this proposal, at the National Cancer Institute. Dr. O'Brien's Laboratory of Genomic Diversity focuses on the coevolution of mammals and retroviruses. Here the principal investigator will continue to develop his skills in molecular biology and reproductive medicine, while learning additional analytic techniques in genomics. The suitability of these research environments and the principal investigator's prior experience makes it feasible to propose to develop new transgenic cat models and stem cell lines to test the specific hypothesis of this proposal: The roles of Oct4 in maintaining pluripotency in Embryonic Stem Cells (ES cells) and the survival of Primordial Germ Cells (PGCs) are conserved in mammals.
In Aim 1, transgenic Oct4-reporter cat embryos will be produced by pronuclear injection to help determine whether Oct4 expression is restricted to pluripotent cells and germ cells in the early embryo and fetus of the cat as in the mouse.
In Aim 2, RNA interference will be used to determine if Oct4 is required for the maintenance of pluripotency in cat embryos and feline ES cells.
In Aim 3, vector mediated transgenesis will be used to test whether Oct4 is required for the survival of feline PGCs. In addition, the potential feline ES and EG cells established in this proposal could be used for future gene-targeting experiments. One of the principal investigator's long-term goals is to realize the potential of the cat as a model species by developing the necessary resources for the research community. By the end of this training program, he will be thoroughly prepared to develop new animal models of human diseases and stem cell and gene therapies.
| Baran, Szczepan W; Johnson, Elizabeth J; Stephens, Matthew A et al. (2009) Development of electronic learning courses for surgical training of animal research personnel. Lab Anim (NY) 38:295-304 |
| Kehler, James S; David, Victor A; Schaffer, Alejandro A et al. (2007) Four independent mutations in the feline fibroblast growth factor 5 gene determine the long-haired phenotype in domestic cats. J Hered 98:555-66 |
| Hubner, Karin; Kehler, James; Scholer, Hans R (2006) Oocytes. Methods Enzymol 418:284-307 |
| Kehler, J; Hubner, K; Scholer, H R (2006) Derivation of germ cells from embryonic stem cells. Ernst Schering Res Found Workshop :125-42 |
| Kehler, James; Hubner, Karen; Garrett, Stacey et al. (2005) Generating oocytes and sperm from embryonic stem cells. Semin Reprod Med 23:222-33 |
