Cathepsin K基因慢病毒质粒载体的构建  

Construction of Cathepsin K Lentivirus Plasmid

作　　者：张艳[1] 柴岗[1] 刘伟[1] 崔磊[1] 曹谊林[1] 

机构地区：[1]上海第二医科大学附属第九人民医院整复外科,200011

出　　处：《组织工程与重建外科杂志》2005年第3期165-166,共2页Journal of Tissue Engineering and Reconstructive Surgery

基　　金：国家重点基础发展规划项目04子项目(G1999054304)。

摘　　要：目的构建Cathepsin K(CTSK)基因慢病毒表达质粒,为进一步研究Cathepsin K在人软骨细胞体外老化过程中的作用奠定基础。方法采用RT-PCR技术扩增Cathepsin K基因的蛋白翻译区,通过连接、转化等分子生物学技术,将该基因克隆至慢病毒pLenti6-V5载体上,经BamH I、XhoI双酶切电泳筛选、鉴定并测序。结果从软骨细胞中成功地克隆出990bp的Cathepsin K基因,并经酶切分析得到6．964Kb和1．005Kb大小的两片断,测序结果显示接头两端序列正确。结论通过基因克隆方法成功构建了Cathepsin K基因表达质粒,可用于进一步的病毒包装,为深入研究该基因对软骨细胞老化的影响奠定了基础。Objective To construct a cathepsin K(CTSK) lentivirus plasmid for exploring the function of cathepsin k in human chondrocytes aging in vitro. Methods RT-PCR was applied to amplify cathepsin k cDNA from the total RNA of human chondrocytes, the cDNA was inserted into plasmid pLenti6/V5-D-TOPO. Results Restricted enzyme analysis and DNA sequencing showed that the sequence of the CTSK lentivirus transgenic plasmid was correct. Conclusion The vector CTSK-lentivirus has been successfully constructed, and it can be used a tool to study the function of cathepsin k.

关 键 词：慢病毒 溶组织蛋白酶K 转基因 

分 类 号：R346[医药卫生—基础医学]