Sox9基因真核表达载体的构建及其诱导大鼠骨髓基质细胞定向分化为骨骺干细胞  

Construction of Sox9 eukaryotic expression vector and its inductive effect on marrow stroma cells differentiating into precartilaginous stem cells

作　　者：胡伟华[1] 陈安民[1] 郭风劲[1] 李锋[1] 黄晖[1] 张伟凯[1] 

机构地区：[1]华中科技大学同济医学院附属同济医院骨科,武汉430030

出　　处：《中国矫形外科杂志》2009年第12期931-934,共4页Orthopedic Journal of China

基　　金：国家自然科学基金资助项目(编号:30650006);教育部博士点基金资助项目[No.教技发中心函(2005)216]

摘　　要：[目的]从永生化骨骺干细胞中克隆Sox9基因并构建真核表达载体,并探讨Sox9诱导骨髓基质细胞向骨骺干细胞分化的可能性。[方法]以RT-PCR方法获得Sox9全长,插入pGEM-TEasy克隆载体中,测序正确后与pEG-FP-IRES2表达载体酶切后连接,复合质粒以脂质体法转染骨髓基质细胞,观察转染效率,Sox9和FGFR-3的表达。流式细胞术鉴定细胞表型,MTT法检测细胞增殖活性。[结果]成功的完成了Sox9的扩增和表达载体的构建,重组载体转染骨髓基质细胞后能检测到Sox9、FGFR-3的表达,增殖活性与骨骺干细胞无异。[结论]成功构建了Sox9真核表达载体,其能诱导骨髓基质细胞分化为骨骺干细胞并具有其特性。[ Objective ] To construct eukaryotic expression vector containing Sox9 derived from immortalized precartilaginous stem cells and to probe its inductive effect on marrow stroma cells differentiating into precartilaginous stem cells. [ Method] Sox9 was obtained with RT - PCR and then inserted into pGEM - T Easy cloning vectors, pEGFP - IRES2 - Sox9 was established after sequencing. Then it＇ s transfected into marrow stroma cells. The expression of Sox9 and FGFR - 3 was detected by Western - blot and immunohistochemisty. Identification and proliferation was determined by flow cytometry and MTT. [ Result] Sox9 cloning and expression vector construction was achieved. Sox9 and FGFR - 3 expression was identified in marrow stroma cells after transfec- tion,whose proliferation activity was similar to precartilaginous stem cells. [ Conclusion ] Sox9 eukaryotic expression vector has been successfully established. Sox9 can induce marrow stroma cells to differentiate into precartilaginous stem cells.

关 键 词：骨骺干细胞 SOX9 骨髓基质细胞 转染 

分 类 号：R346[医药卫生—基础医学]