慢病毒表达载体pLentiTrident1-hBMP2-Neo-hNGF的构建及鉴定  

Construction and identification of a recombinant lentiviral expression plasmid pLentiTrident1-hBMP2-Neo-hNGF

作　　者：韩倩倩[1] 葛少华[1] 闫香珍[1] 杨丕山[1] 

机构地区：[1]山东大学口腔医学院牙周病科山东省口腔生物医学重点实验室,山东济南250012

出　　处：《上海口腔医学》2013年第1期25-29,共5页Shanghai Journal of Stomatology

基　　金：山东省科技发展计划(2010GSF10248);山东大学自主创新基金自由探索项目(2009TS047)~~

摘　　要：目的:构建并鉴定含有人骨形成蛋白2(hBMP2)及神经生长因子(hNGF)目的基因的慢病毒载体。方法:使用限制性内切酶,将新霉素抗性基因(Neo)从载体pLentiTrident1-EGFP-Neo切下,插入pLentiTrident1空载体的相应多克隆位点上,构建出含新霉素抗性的表达载体。采用PCR扩增得到hBMP2与hNGF的目的片段,通过多克隆位点将2个目的基因插入表达载体。对该重组载体进行酶切鉴定、PCR鉴定以及DNA序列测序分析。结果:通过酶切鉴定及测序分析,慢病毒表达载体(plentiTrident 1-hBMP2-Neo-hNGF)构建成功。结论:成功构建了hBMP2与hNGF的共表达慢病毒载体,为研究神经因素在骨组织再生中的作用奠定了基础。PURPOSE： To construct and confirm a recombinant lentiviral vector containing human bone morphogenetic protein 2 （hBMP2） and human nerve growth factor （hNGF）. METHODS： The Neomycin gene was digested from pLentiTridentl-EGFP-Neo and then was subcloned into lentiviral vector. The hBMP2 and hNGF genes were amplified by polymerase chain reaction （PCR）, and then the PCR product was inserted to proper sites of the vector. Finally, the recombinant vector pLentiTridentl-hBMP2-Neo-hNGF was confirmed by agarose gel electrophoresis and DNA sequence analysis. RESULTS： The construction of recombinant lentiviral vector pLentiTrident-hBMP2 -Neo-hNGF was confirmed through restriction enzyme maping analysis and DNA sequencing. CONCLUSIONS： The recombinant lentiviral vector which can coexpress hBMP2 and hNGF is successfully constructed,which lays a solid foundation of studying the effect of neuro factors on bone regeneration. Support by Shandong Provincial Science and Technology Development Plan （2010GSF10248）and Independent Innovation Fund of Shandong University（2009TS047）.

关 键 词：骨形成蛋白2 神经生长因子 慢病毒载体 

分 类 号：R346[医药卫生—基础医学]