LAC REPRESSOR MOUSE

描述(改编自申请者的摘要)：研究目标 在这个应用中提出的是通过控制内源性小鼠基因组 来源于大肠杆菌的乳胶操纵子的成分。它是对 申请者最初提案的目标是获得对 使用这些相同元素的小鼠转基因。即使他们完成了这个目标， 申请者意识到他们努力的最终目标必须是 基因在他们的自然环境中，在那里所有调节和 对活动的影响是存在的。尽管传教士的个人理由是 开发这一系统是为了了解基因调控在 开发性和可塑性，该系统将是一个强有力的工具 了解各种生物和生物的分子基础 生物医学现象。 最初拨款的重点是开发一种能够表达 乳糖抑制物的功能水平，它是细胞内的关键调节蛋白。 小鼠乳胶系统。继续讨论的重点是第二个部分 在该系统中，由抑制物识别的唯一启动子元件，即 Lac运算符。该26个碱基序列的两个拷贝相距约200个碱基 可以赋予调节哺乳动物基因启动子(P53)或 作为转基因被引入时驱动cdna(酪氨酸酶)的哺乳动物启动子 进入小鼠的基因组。下一个明显步骤是引入这些紫胶 通过同源重组进入内源基因启动子的操纵子 (具体目标1)。通过杂交，可以在全球范围内控制目标等位基因 到其中普遍表达Lac抑制子的背景上，例如 作为申请者已经开发的，或者在牢房中-或者 使用限制表达的乳胶抑制物的组织特异性方式 模式(具体目标2)。到项目第二阶段结束时， 目标是将lac系统从转基因小鼠转移到 并开发了一种相对简单的方法学 即使是复杂的表型也可以模拟(具体目标3)。

DESCRIPTION (Adapted from the applicant's abstract): The goal of the research proposed in this application is control of the endogenous mouse genome by elements derived from the lac operon of E. coli. It is a direct extension of the goal of the applicants' original proposal, which was to gain control of murine transgenes using these same elements. Even as they complete this goal, the applicants are aware that the ultimate target of their efforts must be genes in their natural environment, where all the factors that regulate and impinge on activity are present. Although the appicants' personal reason for developing this system is to understand the role of gene regulation in development and plasticity, the system will be a powerful tool for understanding the molecular underpinning of a wide range of biological and biomedical phenomena. The focus of the original grant was the development of a mouse that expresses functional levels of the lac repressor, the key regulatory protein of the murine lac system. The focus of this continuation is on the second component of the system, the unique promoter element recognized by the repressor, the lac operator. Two copies of this 26 bp sequence approximately 200 bp apart can confer the ability to regulate promoters of mammalian genes (p53) or mammalian promoters driving cDNA (tyrosinase) when introduced as transgenes into the murine genome. The next obvious step is to introduce these lac operators into the promoter of an endogenous gene by homologous recombination (Specific Aim #1). Targeted alleles could be controlled globally, by crossing onto a background in which the lac repressor is ubiquitously expressed, such as the one that the applicants have already developed, or in a cell- or tissue-specific manner using lac repressors with restricted expression patterns (Specific Aim #2). By the end of the second phase of the project, the goal is to have transferred the lac system from the transgenic mouse to the endogenous genome and to have developed a relatively simply methodology by which even complex phenotypes can be modeled (Specific Aim #3).