The functional analysis of !6q-linled autosomal dominant cerabeller ataxia associated gene.

本研究的目的是分析与第16染色体长臂相关的易变性相关基因。对于该疾病，迄今为止，我们进行了致病基因座位的鉴定、物理图谱的制作、候选结构域的限定。此外，在puratrophin-1基因中，链式反应非平衡发现了高变化。当我们制备了一种针对puratrophin-1的抗体并在患者大脑中进行化学分析时，该蛋白质与患者小脑浦肯野细胞的细胞骨架蛋白相结合。它是一种基因改变或这是一个病人特有的唯一改变，它是这种疾病的发病的直接原因或它是非常密切相关的。从这样的背景，我们检查它是否这种变化是病因。我们使用来自文库的双杂交系统方法，并克隆与该蛋白质形成相互作用或复合物的蛋白质的cDNA，以检查puratrophin-1基因产物的特征。我们构建了puratrophin-1和酵母转录基因诱饵质粒，将其包含在一起，并将其引入宿主酵母，当我将作为筛选对象的智人胎儿脑cDNA文库连接到酵母转录因子的转录激活结构域并产生它时，我们对在营养培养基上生长的阳性菌落进行了筛选，并从酵母细胞中收集质粒DNA以保持阳性克隆。碱基序列通过直接测序方法解析并鉴定蛋白质，但与细胞骨架蛋白有关的东西并不清楚。我们现在正在研究包括多克隆抗体的制备，原位杂交。可能有这种疾病的其他地方的原因基因，并详细检查候选域，并将其贡献于结果。

A purpose of this study is to analyze it about an associated gene of prepotence to chain the 16th chromosome length arm-related cerebellum transsexualism. As for this disease, we performed identification of a cause gene seat, manufacture of physical map, limitation of a candidate domain till now. Furthermore, in a puratrophin-1 gene, chain reaction non-equilibrium found a high change. When we made an antibody for puratrophin-1 and analyzed it in a patient brain immunohistochemically, the protein cohered with cytoskeletal protein with a patient cerebellum Purkinje cell. It is a gene change or this is an only change peculiar to a patient, and it is a direct cause of the onset of this disease or it was related to extremely closely.From a background such as the above, we inspected it whether this change was etiology. We used two-hybrid system method from a library and did cloning of cDNA of the protein which formed interaction or a complex to this protein to examine a characteristic of a product of a puratrophin-1 gene. We built puratrophin-1 and yeast transcription gene bait plasmid to include and introduced it into host yeast together when I tied the Homo sapiens fetus brain cDNA library which became an object of screening to a transcription activation domain of a yeast transcription factor and produced it. We did screening of the positive colony which we grew on a nutrient medium and collected plasmid DNA from a yeast cell to hold a positive clone in. Base sequence parses this by a direct sequence method and identifies protein, but a thing related to cytoskeletal protein does not become clear. We are examining including manufacture of a polyclonal antibody, in situ hybridization now. There may be a cause gene of this disease elsewhere and examines a candidate domain in detail and is contributing it about the result.