In vivo transfection of foreign genes into the neurons of rat brain as the neurografting method

首先，我研究了控制DNA转染效率的几个因素。我尝试使用质粒DNA作为载体，使用阳离子脂质介导的DNA转染（lipofection）将基因导入小鼠肌肉细胞。通过对脂质素处理DNA的改进，使质粒DNA易于转染。首先，我研究了控制DNA转染效率的几个因素。我尝试使用质粒DNA作为载体，使用阳离子脂质介导的DNA转染（lipofection）将基因导入小鼠肌肉细胞。通过对脂质素处理过的DNA进行改进，质粒DNA可以很容易地通过微注射器注射到小鼠肌肉中，并且可以被注射点附近的肌肉细胞掺入和表达。形态学上，我使用了两种质粒DNA， l7rh - β -gal和prsvl - β -gal来检测转染后的DNA在小鼠肌肉细胞中的表达。l7rh - β -半乳糖质粒DNA含有来自SV40 t抗原基因和大肠杆菌- β -半乳糖苷酶基因的不明确定位信号，这两个基因都融合到SV40病毒增强启动子上。当用l7rh质粒DNA转染肌肉细胞时，所产生的大肠杆菌β -半乳糖苷酶蛋白可以运输到细胞核中，因此细胞核应密集地用X-gal染色，呈圆形。相反，使用prsvl - β -gal，光镜下观察到一些x -gal染色的小鼠肌肉纤维束呈蓝色肌肉。电镜观察发现，X-gal染色后的肌肉肌膜中有大量精细的电子致密物质。虽然我们还没有发现任何控制脂质体dna对小鼠肌肉细胞效率的因素，但这项技术的改进将为我们从形态和功能方面研究基因在特定肌肉细胞中的作用提供有力的工具。下一阶段，我打算将此方法应用于神经移植。少

First of all, I investigated the several factors controlling the efficiency of DNA transfection. I tried to use plasmid DNA as a vector for the introduction of genes into mouse muscles cells using cationic lipid-mediated DNA transfection(lipofection).By this improvement of this lipofectin-treated DNA, plasmid DNA were easily First of all, I investigated the several factors controlling the efficiency of DNA transfection. I tried to use plasmid DNA as a vector for the introduction of genes into mouse muscles cells using cationic lipid-mediated DNA transfection(lipofection).By this improvement of this lipofectin-treated DNA, plasmid DNA were easily injected into the mouse muscles through a microsyringe, and could be incorporated and expressed by muscle cells in the vicinity of injection point Morphologically I used two kinds of plasmid DNAs, L7RH-beta-gal and pRSVL-beta-gal, in order to detect the expression of transfected DNAs in mouse muscles cells. The L7RH-beta-gal plasmid DNA contain … More s the unclear location signal from the SV40 T-antigen gene and E.coli-beta-galactosidase gene both of which are fused to the SV40 virus enhancer-promoter. Since the produced E.coli beta-galactosidase proteins can be transported into nuclei of cells when the muscle cells were transfected with the L7RH-plasmid DNA, the nuclei should be densely stained with X-gal in a round shape. On the contrary, using the pRSVL-beta-gal, several number of the X-gal-stained mouse muscles fiber bundles were observed as blue colored muscles by light microscopy. By electron microscopic investigation, the large number of fine electron dense materials were occupied in the sarcolemma of the X-gal stained muscles.Although we have not identified any factors controlling the efficiency of lipofectin to mouse muscles cells with plasmid DNAs, improvements of this technique would provide us with a powerful tool for studying gene action in particular muscle cells in relation to morphological and functional aspects.Next stage, I will intend to apply this procedure to the neurografting. Less