大鼠心肌H^+-ATP酶抑制蛋白基因克隆及初步鉴定  

Cloning of cDNA of inhibitory protein of rat myocardium H +-ATPase

作　　者：朱世军[1] 李芳[1] 陈伟[1] 姜晓姝[1] 徐曼[1] 徐长庆[1] 王孝铭[1] 陈忠斌 于康震 

机构地区：[1]哈尔滨医科大学病理生理教研室 [2]哈尔滨医科大学遗传教研室

出　　处：《中国病理生理杂志》1998年第2期153-156,共4页Chinese Journal of Pathophysiology

摘　　要：目的和方法：根据大鼠线粒体Ｈ＋－ＡＴＰ酶抑制蛋白基因（ＩＦ１）ｃＤＮＡ序列，设计并人工合成一对特异性扩增ＩＦ１ｃＤＮＡ的引物。以提取的大鼠心肌ｍＲＮＡ为模板，反转录－聚合酶链式反应扩增出约４００ｂｐ条带，以Ｋｌｅｎｏｗ酶处理、纯化ＰＣＲ产物，与经Ｓｍａｌ酶切线性化的ｐＵＣ１９质粒ＤＮＡ连接，经转化、筛选初步获得两个重组质粒。对重组质粒进行酶切图谱分析并以重组质粒为模板做ＰＣＲ检测。结果：该质粒为ＩＦ１ｃＤＮＡ基因重组ｐＵＣ１９，并且该基因以正向插入，命名为ｐＩＦ１。AIM and METHODS:A pair of primers were designed and synthesized based on ATPase inhibitory protein cDNA sequences of rat myocardium mitochondrial H +-ATPase. A 400bp cDNA fragment was amplified by reverse transcription-polymerase chain reaction(RT-PCR) using mRNA of rat myocardium as template.IF 1 cDNA was blunted with Klenow enzyme and purified using geneclean kit,the PCR amplified cDNA was inserted into pUC19 at Sma Ⅰ site and the recombinant plasmid DNA was used to transform the competent E.coli DH 5α cells.Recombinant plasmids were picked up and miniprepered. RESULTS:A clone,pIF 1,bearing IF 1 cDNA was identified by restriction analysis and PCR amplification using recombinant pUC19 as template.The IF 1 cDNA in the plasmid was found in right orientation by restriction analysis.

关 键 词：大鼠 心肌缺血 腺苷三磷酸酶类 基因克隆 CDNA 序列分析 

分 类 号：R542.2[医药卫生—心血管疾病]