TARGETED MUTATIONS OF TYROSINE KINASE GENES IN THE MOUSE

该项目将包括选择方法的开发 用于将逆转录病毒DNA整合到特定宿主基因中 产生特定的突变；并针对逆转录病毒感染 到含有特定细胞表面蛋白的特定细胞类型。 在我们之前的工作中，我们已经证明了 通过插入失活X连锁基因产生突变 小鼠编码次黄嘌呤-鸟嘌呤磷酸核糖转移酶基因 (HGPRTase)。我们建议扩大这些研究的范围，以促进 干扰其他更有趣的靶基因，并改进 这个过程的效率。我们将尝试选择 隐性抗癌基因中断所致的转化子， 最近为视网膜母细胞瘤系统演示了。另一个 强大的选择是细胞诱导后的存活 末端分化。我们还将尝试选择 将逆转录病毒基因组插入特别受调控的基因 方法：这一过程类似于使用的Mud-lac基因融合 在细菌中。最后，我们描述了为提高效率所做的努力 靶向特定宿主基因的逆转录病毒DNA的研究 整合到病毒基因组中的潜在区域 与目标基因同源。在一个单独的子项目中，我们将 尝试将病毒定位于特定类型的细胞。在其中的一些中 实验表明，病毒的env基因会被部分破坏， 被编码金黄色葡萄球菌蛋白的基因取代 A.将这些病毒暴露于特定的单抗免疫球蛋白 制剂将用抗体包裹病毒，定位于 变量特异性--向外决定结构域。的能力 这些病毒进入并生长在表达抗原的细胞上 被识别的单抗将被检测通过测量 感染后产生的子代病毒。

This project will consist of the development of methods to select for the integration of retroviral DNAs into particular host genes to generate specific mutations; and to target retroviral infection to particular cell types bearing specific cell-surface proteins. In our previous work, we have demonstrated the feasibility of generating mutations by insertional inactivation of the X-linked mouse gene encoding hypoxanthine-guanine phosphoribosyl transferase (HGPRTase). We propose to extend these studies to facilitate the disruption of other more interesting target genes, and to improve the efficiency of the process. We will attempt to select for transformants due to the disruption of recessive anti-oncogenes, recently demonstrated for the retinoblastoma system. Another powerful selection is survival of cells after induction toward terminal differentiation. We will also attempt to select for insertion of retroviral genomes into genes regulated in particular ways; this procedure is analogous to the Mud-lac gene fusions used in bacteria. Lastly, we describe efforts to improve the efficiency of the targeting of retroviral DNAs to particular host genes by the incorporation into the viral genome of potential regions of homology with the target gene. In a separate subproject, we will try to target virus to particular cell types. In some of these experiments the viral env gene will be partially disrupted and substituted with the gene encoding the Staphlococcus aureus protein A. Exposure of these virus to particular monoclonal IgG preparations will coat the virus with antibody, oriented with the variable specificity-determining domain outward. The ability of these viruses to enter and grow on cells expressing the antigen recognized by the monoclonal antibody will be tested by measuring the production of progeny virus after infection.