Use of Xenopus Oocyte/mRNA Injection System for the Study of Plant Hormone Receptors

非洲爪蟾卵母细胞/mRNA注射系统提供了一个关键的 各种动物cDNA鉴定和克隆的关键 神经递质受体 萨顿博士建议这个系统 可以类似的方式用于植物激素的研究 （植物激素）受体。 这项建议的具体目的是 测试注入非洲爪蟾卵母细胞的植物mRNA 将导致植物激素的合成和掺入 受体进入非洲爪蟾卵母细胞质膜，这些 受体将与非洲爪蟾卵母细胞的第二信使 系统. 将从大麦糊粉层中分离RNA， 对各种植物激素都有反应 RNA将 注射到非洲爪蟾卵母细胞中， 培养3-5天。 成功的合成和整合 受体进入卵母细胞膜的能力将通过以下方法进行测试： 监测植物激素诱导的氯电流， 双电极电压钳技术 技术和放射性标记的激素结合研究。 如果这种方法成功，这项研究的结果将 促进植物激素的克隆和表征 受体已经被证明是难以捉摸的更传统的生化 接近。 植物激素的表达或缺乏表达 这种动物细胞系统中的受体也应该提供 关于蛋白质分选到血浆的重要信息 膜的

The Xenopus oocyte/mRNA injection system has provided a critical key to the characterization and cloning of cDNAs for various animal neurotransmitter receptors. Dr. Sutton proposes that this system can be used in an analogous manner for the study of plant hormone (phytohormone) receptors. The specific aim of this proposal is to test the hypothesis that plant mRNA injected into Xenopus oocytes will result in the synthesis and incorporation of phytohormone receptors into the Xenopus oocyte plasma membrane, and that these receptors will couple to the Xenopus oocyte second messenger systems. RNA will be isolated from the barley aleurone layer which is known to be responsive to various phytohormones. The RNA will be injected into Xenopus oocytes and the oocytes allowed to incubate for 3-5 days. The successful synthesis and incorporation of the receptors into the oocyte membrane will be tested by monitoring for phytohormone-induced chloride currents in the membrane of the oocytes by the two-electrode voltage clamp technique and by radio-labelled hormone binding studies. If this approach is successful, the results of this research will facilitate the cloning and characterization of plant hormone receptors which have proven elusive to more traditional biochemical approaches. The expression or lack of expression of plant hormone receptors in this animal cell system should also provide significant information about protein sorting to the plasma membrane.