| Budget Amount *help |
¥3,800,000 (Direct Cost: ¥3,800,000)
Fiscal Year 2004: ¥1,700,000 (Direct Cost: ¥1,700,000)
Fiscal Year 2003: ¥2,100,000 (Direct Cost: ¥2,100,000)
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| Research Abstract |
The selective reaction to the gene would have the potential for site-directed mutation. We already demonstrated that triple helix oligonucelotides (TFO) bearing 2-amino-6-vinypurine derivative achieved the selective point mutaitions to a target site in a shuttle vector plasmid, which replicates in mammalian cells. These reactions were very effective in vitro under acidic conditions, but the reactive derivatives to react to the target under neutral condition are necessary for application to the cell. In 2003,we have investigated the new triplex mediated cross-linking reagents with high selectivity toward the target under neutral conditions. We synthesized those reactive TFO using post modification method and studied about the reactivity of them. Unfortunately, these reactive TFO did not react to the target duplex, because these oligonucelotides having reactive molecules were not be able to form the stable triple helix with the targets. Thus, in 2004, we have attempted to develop the novel more reactive derivatives. In these results, we have exploited the more reactive derivatives, which react to cytidine selectively to give the adducts in 50 % yield only after 5 hrs. In addition, we have demonstrated that reactive oligonucelotides increase the efficiency of the antisense inhibition in the cell. These results suggest that these cross-linking reagents may react with the target in the cell. In conclusion, we have developed the high effective cross-linking reagents to react in the cell. Based on the results, we would like to investigate the development of chemical methods to occur the point mutation in the cell.
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