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目的:建立AM L1-ETO融合基因转基因小鼠,在整体动物水平研究AM L1-ETO融合蛋白在白血病发病中所起的作用。方法:构建hCG/AM L1-ETO转基因质粒,通过显微注射将该质粒转入小鼠受精卵中,植入假孕母鼠输卵管,获得G0代转基因小鼠;用聚合酶链反应(PCR)方法检测融合基因的整合情况,逆转录-聚合酶链反应(RT-PCR)方法检测融合基因的表达情况和组织表达谱。结果:PCR法共检出10只G0代转基因阳性小鼠,其中8个系均产下F1代小鼠,由此建立了8个AM L1-ETO转基因小鼠系。RT-PCR法证实AM L1-ETO融合基因在22系中稳定表达,但血常规、肝脏、脾脏等组织未见异常变化;对该系小鼠的组织表达谱检测表明,融合基因在肝脏、脾脏、心脏和肌肉组织存在不同程度的表达,但在脑组织中不表达。结论:AM L1-ETO融合基因能在转基因小鼠的骨髓等组织中表达,但尚不足以引发白血病,推测其他致病基因在小鼠白血病的发生中也起一定作用。
OBJECTIVE: To establish AM L1-ETO fusion gene transgenic mice to study the role of AM L1-ETO fusion protein in the pathogenesis of leukemia at the whole animal level. Methods: The hCG / AM L1-ETO transgenic plasmid was constructed and transfected into mouse fertilized eggs by microinjection. The pseudopregnant ovariectomized mice were implanted into the oviducts to obtain G0 transgenic mice. Polymerase chain reaction (PCR) Methods The integration of fusion gene was detected by RT-PCR and the expression of fusion gene and tissue expression pattern were detected by reverse transcription-polymerase chain reaction (RT-PCR). Results: Ten G0 transgenic mice were detected by PCR, of which 8 were all F1 generation mice, and 8 AM L1-ETO transgenic mice lines were established. RT-PCR confirmed the stable expression of AM L1-ETO fusion gene in 22 lines, but no abnormal changes were found in blood, liver, spleen and other tissues. Tissue expression profiling of the mice showed that the fusion gene was expressed in liver and spleen , Heart and muscle tissue to varying degrees of expression, but not expressed in brain tissue. CONCLUSION: AM L1-ETO fusion gene can be expressed in bone marrow and other tissues of transgenic mice but not enough to cause leukemia. It is speculated that other pathogenic genes play a role in the development of mouse leukemia.