Mouse spinal marrow in-vitro electrictransfection gene method, mouse spinal marrow electrictransfection tissue slice culture model establishing method and application thereof
A model building and transgenic technology, applied in the field of nervous system gene function research, can solve problems such as the difficulty of transferring foreign genes into the spinal cord, achieve the effect of reducing the difficulty of operation and ensuring no necrosis
Inactive Publication Date: 2018-11-20
XINXIANG MEDICAL UNIV
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Problems solved by technology
But so far, there has been no research on electrotransfer in the mouse spinal cord. Because it is very difficult to accurately inject exogenous genes into the spinal cord cavity of the embryo in the mouse uterus, it is difficult to transfer exogenous genes into the spinal cord. The study of the influence of the source gene on neurons in the spinal cord during mouse embryonic development has also encountered great difficulties
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Embodiment 1
[0031] Materials and instruments used in this embodiment are as follows:
[0032] The reporter gene pCAGGS-GFP plasmid (green fluorescent protein) was donated by Dr. Redies, and was amplified and extracted in the laboratory; the pregnant mice used in the research were C57 mice raised in the laboratory; the plasmid DNA extraction kit (Kangwei Century Biotechnology Co., Ltd. ); 4'6-diamidino-2-phenylindole (DAPI, Roche); Neurobasal medium (Invitrogen, USA); hanging cell culture dish (PICM03050, Millipore, USA); SZM45 stereo dissecting microscope ( Zhejiang Sunny Instrument Co., Ltd.); CUY-21 (BEX) multifunctional in vivo electroporation instrument (NEPA company in Japan); M205FA inverted stereo microscope (Leica company in Germany); VT1200S oscillating microtome (Leica company in Germany); P- 2000 Needle Pulling Instrument (Sutter, USA); Confocal Laser Microscope (Olympus ix81, Olympus Corporation, Japan).
[0033] The preparation of main reagent in the present embodiment is as...
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Abstract
The invention relates to a mouse spinal marrow in-vitro electrictransfection gene method, a mouse spinal marrow electrictransfection tissue slice culture model establishing method and application thereof, and belongs to the technical field of research on a nervous system gene function. The mouse spinal marrow in-vitro electrictransfection gene method comprises the following steps: taking an embryonic mouse out of the uterus of a mother mouse, stripping the spinal marrow out of the embryonic mouse, injecting plasmid containing a target gene into a spinal marrow cavity of the in-vitro spinal marrow, putting an electrode of an electrictransfection instrument on the two sides of the spinal marrow, putting a positive electrode on one side needing electrictransfection, and performing electrictransfection to obtain electrictransfection spinal marrow. The electrictransfection spinal marrow is subjected to tissue slice culture, a mouse spinal marrow electrictransfection tissue slice culture model can be established by the method, and a new research method is provided for the influence on neuron in the spinal marrow by an exogenous gene in the development of the mouse embryo, so that effective means for treating the spinal marrow diseases is found; and the method is more suitable for dynamic change observation of the neuron and is a research technology between a living body and a cell.
Description
technical field [0001] The invention relates to a mouse spinal cord in vitro electrical transgene method, a mouse spinal cord electrical transfer tissue slice culture model establishment method and an application thereof, belonging to the technical field of nervous system gene function research. Background technique [0002] Mammalian embryonic development is carried out in the mother's body, so it is difficult to study the abnormalities of the ganglion system during embryonic development. Mice are a commonly used experimental model animal. In the previous research process, we have carried out the mouse intrauterine electrotransduction technology in the mouse model. This technology realizes the abnormality of exogenous genes in the early cerebral cortex of mouse embryos expression, and analyze its effect on the cerebral cortex at a specific stage (Yang Ciqing, Li Xiaoying, Lu Xi, et al. Establishment of an in utero electrical transgenic method in mice[J]. Journal of Anatomy,...
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IPC IPC(8): C12N15/85C12N15/65C12N5/10
CPCC12N5/0618C12N15/65C12N15/85C12N2510/00C12N2533/76C12N2800/107
Inventor 杨慈清林俊堂李小英李栓庆管丽红乔梁李晗
Owner XINXIANG MEDICAL UNIV