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目的 制备HLA B2 70 4转基因小鼠。方法 应用显微注射将HLA B2 70 4基因注入C5 7BL/6×昆明鼠和昆明鼠×昆明鼠F1代受精卵 ,出生动物及其后代经PCR初筛 ,采用斑点杂交和Southern杂交对阳性标本作进一步鉴定和整合基因拷贝数测定 ,阳性者行免疫组化染色和流式细胞术检测HLA B2 70 4蛋白表达水平。结果 10只仔鼠 (首建转基因鼠 ,founder)表达有HLA 2 70 4基因。由注射基因后移卵出生的 10 1只小鼠中 ,C5 7BL/6×昆明鼠杂种 3 7只 ,阳性 7只 ( 18 9% ) ,昆明鼠×昆明鼠杂种 64只 ,阳性 3只 ( 4 7% ) ,阳性动物外周血淋巴细胞、皮肤和胃肠道上皮细胞内均有HLA B2 70 4蛋白表达。结论 应用C5 7BL/6和昆明鼠成功建立了HLA B2 70 4的转基因小鼠。与昆明鼠×昆明鼠F1代相比 ,C5 7BL/6×昆明鼠F1代更适合用于转基因动物制备。
Objective To prepare HLA B2 70 4 transgenic mice. Methods HLA B2 70 4 gene was injected into the F1 generation of C5 7BL / 6 × Kunming mice and Kunming mice × Kunming mice by microinjection. The animals and their progenies were preliminarily screened by PCR. The positive samples were identified by dot blot and Southern hybridization Further identification and integration of gene copy number determination, positive by immunohistochemical staining and flow cytometry HLA B2 70 4 protein expression levels. Results 10 offspring mice (founder transgenic mice, founder) expressed HLA 2 70 4 gene. Among 10 1 mice born after transgene injections, 37 were C5 7BL / 6 × Kunming mice, 7 were positive (18 9%), 64 were Kunming mice × Kunming mice and 3 were positive (4 7%), positive animals, peripheral blood lymphocytes, skin and gastrointestinal epithelial cells have HLA B2 70 4 protein expression. Conclusion The transgenic mice of HLA B2 70 4 were successfully established by using C5 7BL / 6 and Kunming mice. Compared with Kunming mice × Kunming mice F1 generation, F1 generations of C5 7BL / 6 × Kunming mice are more suitable for the preparation of transgenic animals.