L1 mutagenesis for mammalian models of human diseases

描述(申请人提供)：转基因公司正在开发一种创新的哺乳动物随机突变系统，该系统使用L1反转录转座子直接进行生殖系突变。基因工程公司的目标是培育出“突变大鼠”。在这些大鼠中，突变将发生在精子中，一些后代将包含单基因中断。我们将使用小鼠来优化和验证我们的系统，因为生产转基因大鼠的成本要高得多。我们已经培育出含有标记的L1转基因的“突变小鼠”，该转基因能够进行胚系插入突变。这项建议的重点是验证转基因随机突变系统作为功能基因组学工具的有效性。该项目的第一阶段包括突变小鼠的特征。具体目的1是确定突变小鼠精子中插入突变的频率。具体目标2是确定插入到基因中的插入片段的百分比。具体目标3是确定基因破坏的效率。该项目的第二阶段将使用从第一阶段小鼠研究中获得的信息来创造突变大鼠。L1元件存在于所有哺乳动物中，人类的L1元件能够在人、小鼠、仓鼠、猫科动物甚至是鹌鹑细胞中进行高水平的逆转录转座。因此，我们在小鼠身上的研究结果应该直接推论到大鼠。转基因的随机突变系统完全可以用于任何哺乳动物。突变小鼠和大鼠将通过为新药靶点建立动物模型，为药物发现计划提供实质性的帮助。

DESCRIPTION (provided by applicant): Transposagen is developing an innovative mammalian random mutagenesis system that uses an L1 retrotransposon for direct germ-line mutagenesis. Transposagen's goal is to produce "mutator rats". In these rats, mutagensis will occur in sperm and some offspring will contain single gene disruptions. We will use the mouse to optimize and validate our system because of the much higher cost in producing transgenic rats. We have developed "mutator mice" containing a marked L1 transgene capable of germ-line insertional mutagenesis. The focus of this proposal is to validate the Transposagen random mutagenesis system as a tool for functional genomics. Phase I of this project includes characterization of the mutator mice. Specific aim 1 is to determine the frequency of insertional mutagenesis in the sperm of mutator mice. Specific aim 2 is to determine the percentage of insertions that insert into genes. Specific aim 3 is to determine the efficiency of gene disruption. Phase II of the project will use the information gained from the Phase I mouse studies to create mutator rats. L1 elements are present in all mammals and a human L1 element is capable of high-level retrotransposition in human, mouse, hamster, feline, and even quail cells. Therefore, our findings in mice from this study should extrapolate directly to rats. Transposagen's random mutagenesis system could utlimately be adapted for use in any mammal. Mutator mice and rats will substantially aid drug discovery programs by producing animal models for new drug targets.