| Research Abstract |
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
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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
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