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自噬基因BECN1真核表达载体和miRNA表达载体构建

发表时间:2011-09-15  浏览量:1341  下载量:636
全部作者: 任以中,李楚芳,潘蔚琦,孙彩军,陈凌
作者单位: 中国科学院广州生物医药与健康研究院,呼吸疾病国家重点实验室;中国科学技术大学生命科学学院
摘 要: 以人cDNA为模板,根据已知BECN1基因序列设计引物,通过PCR成功扩增出BECN1基因编码序列,并连接至真核表达载体pCDNA4上。根据PCR和酶切鉴定,克隆构建成功。研究还设计了针对BECN1编码区的特异miRNA,并成功构建出表达该特异miRNA的表达载体pCDNA6.2-miR-beclin1. 通过Western blotting检测,所构建的真核表达载体pCDNA4-beclin1成功表达BECN1蛋白,并可以被特异miRNA表达载体pCDNA6.2-miR-beclin1干扰。研究构建的BECN1真核表达载体和miRNA表达载体为进一步研究BECN1蛋白的生物学功能奠定了基础。
关 键 词: 细胞生物学;BECN1;真核表达载体;miRNA表达载体
Title: Construction of eukaryotic expression and miRNA expression vector of autophagy gene BECN1
Author: REN Yizhong, LI Chufang, PAN Weiqi, SUN Caijun, CHEN Ling
Organization: State Key Laboratory of Respiratory Disease, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences; School of Life Sciences, University of Science and Technology of China
Abstract: The coding sequence of BECN1 was obtained from human cDNA library by specific primers using PCR and was cloned to eukaryotic expression vector pCDNA4. The construct was verified by PCR and enzymatic digestion. Specific miRNA targeting BECN1 ORF was designed and cloned to miRNA expression vector pCDNA6.2. The eukaryotic expression vector could express BECN1 protein and was interfered by the specific miRNA expression vector. The vectors provided a good tool to study the biological function of BECN1.
Key words: cell biology; BECN1; eukaryotic expression vector ; miRNA expression vector
发表期数: 2011年9月第17期
引用格式: 任以中,李楚芳,潘蔚琦,等. 自噬基因BECN1真核表达载体和miRNA表达载体构建[J]. 中国科技论文在线精品论文,2011,4(17):1556-1561.
 
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