Mouse model of human diseases caused by mtDNA mutations

mtDNA突变引起的人类疾病的小鼠模型

• 批准号: 6962761
• 负责人: MICHAEL D KOOB
• 金额: $ 16.58万
• 依托单位: UNIVERSITY OF MINNESOTA
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-07-15 至 2007-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6962761
• 关键词: disease /disorder model; fungal genetics; gene mutation; genome; green fluorescent proteins; mitochondrial DNA; model design /development; polymerase chain reaction; tissue /cell culture; yeasts

DESCRIPTION (provided by applicant): Mouse models of human genetic diseases have been critical for confirming that particular mutations cause disease, for obtaining a detailed molecular understanding of the pathophysiology of these diseases, and for developing and testing potential therapies. Technical limitations, however, currently prevent us from generating accurate mouse models of human diseases caused by mutations in our mitochondria! DNA. Although several groups have shown that they can generate mice that contain mitochondrial genomes isolated from mouse tissue culture cells, the technology has not yet been developed that allows us to introduce specific nucleotide changes into mouse mitochondrial genomes. We are proposing to overcome this technical limitation. We have cloned the complete mouse mitochondrial genome stably in E. coli and have shown that we can introduce essentially any desired mutation into these clones. We have also shown that we can introduce these engineered mitochondrial genomes into the mitochondria of yeast cells devoid of their own mtDNA and that these mouse mtDNA genomes are replicated accurately in these cells. We now propose to use these transgenomic yeast mitochondria as vectors for transferring the engineered mtDNA genomes back into the mitochondrial networks of mouse tissue culture cells, which will then be used to generate mice with modified mitochondrial genomes. Our specific aims for this project are to 1) develop and optimize methods for using transgenomic yeast mitochondria as mitochondrial delivery vectors that will efficiently fuse to mouse mitochondrial networks, and to 2) determine how to best screen or select for mouse cells that have been transformed with modified mitochondrial genomes.

描述（由申请人提供）：人类遗传疾病的小鼠模型对于确认特定突变导致疾病、获得这些疾病的病理生理学的详细分子理解以及开发和测试潜在疗法至关重要。然而，由于技术上的限制，目前我们还无法建立精确的小鼠模型，来研究线粒体突变引起的人类疾病。DNA.尽管有几个研究小组已经证明，他们可以产生含有从小鼠组织培养细胞中分离的线粒体基因组的小鼠，但尚未开发出允许我们将特定核苷酸改变引入小鼠线粒体基因组的技术。我们建议克服这一技术限制。我们已经在大肠杆菌中稳定地克隆了小鼠线粒体基因组的全基因组。大肠杆菌，并已表明，我们可以引入基本上任何所需的突变到这些克隆。我们还表明，我们可以将这些工程化的线粒体基因组引入缺乏自身mtDNA的酵母细胞的线粒体中，并且这些小鼠mtDNA基因组在这些细胞中准确复制。我们现在建议使用这些转基因酵母线粒体作为载体，将工程化的mtDNA基因组转移回小鼠组织培养细胞的线粒体网络中，然后将其用于产生具有修饰的线粒体基因组的小鼠。我们这个项目的具体目标是1）开发和优化使用转基因酵母线粒体作为线粒体递送载体的方法，这些载体将有效地融合到小鼠线粒体网络中，以及2）确定如何最好地筛选或选择已经用修饰的线粒体基因组转化的小鼠细胞。