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Show posters on the hill II A New Method of Gene Targeting Aubrey Chan and Professor Erik Jorgensen Departments of Bioengineering and Biology ie soil nematode Caenorltabdttis elegans: a simple and easy-to-use animal. Gene targeting allows for specific changes to a specific gene, opening many possibilities for both therapy and research. 1 1 I 1 JXLL___ ^^^1 Homologous recombination is a rurally ^^™ repair broken DNA. With some . waxing, hum ever, we f may be able to get it to v vork for us. -' ¦>¦¦ working with unc-119, a gene responsible for stabilizing rvous system. In humans, failures in this function have nked to such catastrophic events as grand mal seizures. We are interested in the genes that allow nerves to work - genes that allow them to grow, connect, regenerate, and transmit signals in the body. Instead of studying these genes in an animal like a lab mouse, we are study-ing them in the small soil worm, C. elegans. These animals are tiny but have nerves that work in the same way as mouse or human nerves. In addition, worms grow up in only a few days, are transparent (so we can see the nerves from the outside), and can be grown in vast quantities at low cost. One of the best ways to study and understand genes' molecular functions is to mutate the genes and observe the effects. This process is like figuring out how a complicated machine works by removing one part at a time and seeing how the machine fails when that part is missing. Since the early seventies, researchers have been using molecular sledgehammers to knock off genes one at a time to discover their functions. Ideally, however, a researcher would be able to mutate a gene of interest) and only the gene of interest) in a controlled way to study the effects of specific changes to the DNA sequence - a molecular screwdriver with great precision. Dr. Mario Capecchi at the University of Utah has developed such techniques for making specific gene knockouts in mice. No one, however, has yet found a way to do these knockouts in worms. We are developing a technique to induce such specific mutations by hijacking the cell's natural DNA repair machinery. To accomplish this task, we will engineer a unique endonuclease, an enzyme designed to recognize and cut specifically the gene we want to mutate, and use a customized, "donor" version of our gene of interest that has the new sequence we want the cell to have. The first gene we are using for this work is called unc-119, a gene im-plicated in nervous system stability. When all the components meet under the right conditions, cuts will occur on both the native and donor genes, in different places. When the cell detects this broken DNA, its repair machinery will go to work, hopefully repairing the breaks in such a way as to incorporate our donor DNA, thus mutating the native chromosome. Strong selection for worms mutated in such a way will serve to increase the yield of success-fully mutated worms. This technique has been shown to work at high frequency in the fruit fly D. melanogaster. {68} |