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人纤溶酶原激活物抑制物1转基因小鼠的建立

来源:论文汇编 作者:自动采集 2005-1-1
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摘要: 摘要:目的建立人纤溶酶原激活物抑制物1(PAI-1)转基因小鼠。方法利用构建的高效真核表达载体pCAGGS—PAI-1,制备注射用DNA。以显微注射法将目的DNA注入小鼠受精卵,再将受精卵移植到受体母鼠,制备转基因小鼠。经鼠尾组织DNA提取、PCR筛选、Southern杂交、DNA序列测定对转基因小鼠进行鉴定。...



  摘 要:目的建立人纤溶酶原激活物抑制物1(PAI-1)转基因小鼠。方法利用构建的高效真核表达载体pCAGGS—PAI-1,制备注射用DNA。以显微注射法将目的DNA注入小鼠受精卵,再将受精卵移植到受体母鼠,制备转基因小鼠。经鼠尾组织DNA提取、PCR筛选、Southern杂交、DNA序列测定对转基因小鼠进行鉴定。结果对获得的29只小鼠经PCR筛选、Southern杂交分析、DNA序列测定,得到了2只整合了人PAI-1cDNA的转基因小鼠。在刚出生的转基因小鼠尾部及后肢发现出血性改变,免疫组织化学观察到在,肾脏局部PAI-1表达稍有增高。结论本研究中,我们建立了人PAI-1基因转基因小鼠,为进一步研究PAI-1在肾脏疾病中调控细胞外基质降解的机制提供了动物模型,为研究利用抗凝、促纤溶疗法治疗慢性肾炎奠定了基础。

  Construction of mice transgenic for human plasminogen-activator inhibitor type 1

  LIU Shuwen ,CHEN Xiangmei ,Ye Yizhou

  (Division of Nephrology,General Hospital of PLA,Key Laboratory and Kidney Center of PLA,Beijing 100853,China)

  Abstract:Objective To construct mice transgenic for human plasminogen-activator inhibitor type 1 (PAI-1),and to probe the role and mechanism of PAI-1 in the synthesis and degradation of extracellular matrix in kidney diseases.Methods The eukaryotic expression plasmid pCAGGS-PAI-1 was digested with Pvu Ⅰ and Hind Ⅲ,recovered using agarose-gel electrophoresis,and then purified by using CsCl density-gradient ultracentrifugation.DNA was microinjected into male pronuclei of fertilized mouse eggs,the injected mouse eggs were then transplanted into the oviduct of pseudopregnant female mice.The offsprings were screened by using polymerase-chain reaction,and then identified by Southern-blot analysis and DNA sequencing.Results Two of twenty-nine survived offsprings were identified to be mice transgenic for human PAI-1.The mice transgenic showed subcutaneous hemorrhage at the end of the tail and hind limb.Immunohistochemical examination showed slightly elevated PAI-1 expression in the kidney.Conclusion Mice transgenic for human PAI-1 were constructed,providing an animal model for further investigations of the role and mechanism of PAI-1 in the pathogenesis and progression of kidney diseases.

  Keywords:Plasminogen-activator inhibitor type 1;Transgenic mice

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