人牙本质磷蛋白功能片段的真核表达  

Eukaryotic expression of human dentin phosphoprotein domain

作　　者：万领[1] 李玉晶[1] 张福萍[2] 张瑾[2] 周伟[2] 董小平[2] 

机构地区：[1]首都医科大学附属北京口腔医院,北京100050 [2]中国疾病预防控制中心病毒病预防控制所

出　　处：《现代口腔医学杂志》2005年第3期262-265,共4页Journal of Modern Stomatology

摘　　要：目的　本研究旨在利用分子生物学技术真核表达包含功能区的人DPP片段。方法　构建了人DPP蛋白5′端基因片段的杆状病毒表达质粒pFastBacHb -hDPP -N ,经BH1 0BAC转座反应后感染昆虫细胞sf9,进行重组蛋白的表达。结果　经2～3代后,重组病毒产生明显的CPE ,人DSPP蛋白特异性多克隆抗体Westernblot鉴定,结果显示重组病毒感染3代毒sf9细胞的裂解物中在约1 7KD位置上出现了特异性的反应条带。结论　表明杆状病毒系统可表达人DPP基因片段。为进一步通过对人DPP功能片段的研究揭示DPP在牙齿发育和牙髓修复中的作用及其机制奠定基础。Objective In this study, the human Dentin Phosphoprotein(DPP) domain containing DSS was cloned and expressed by using genetic engineering in eukaryotic cells with Bac-to-Bac bacular expression system.Methods The 504-bp fragment of human DPP gene was cloned into the the donor plasmid pFastBacHb. After transposition with BH10BAC, recombinant Bacmid DNAs were isolated and transfected in Sf9 cells.Results The typical cytopathologic effect caused by the recombinant baculovirus became obvious, after two or three passages. The expression of DPP-derived protein was screened by Western blot assays with a polyclonal antibody specific for human DSPP. It revealed a positive band mobilizing around 17 KD in the lysate of SF9 cells.Conclusion It suggests that the N-terminus DPP peptide could be expressed in the baculovirus expression system. It will lay a theoretical foundation for revealing the mechanism of DPP on tooth development and the effect on pulp restoration.

关 键 词：真核表达 功能片段 牙本质磷蛋白 分子生物学技术 昆虫细胞SF9 Western 杆状病毒系统 基因片段 多克隆抗体 sf9细胞 DPP 表达质粒 重组蛋白 重组病毒 blot 病毒感染 牙髓修复 牙齿发育 P蛋白 特异性 功能区 CPE DSP 裂解物 

分 类 号：R780.2[医药卫生—口腔医学]