人生长激素的体外重组及在原代鼠成肌细胞内的表达  

Construction of vector encoding human growth hormone and expression in murine primary myoblasts

作　　者：李锋[1] 卢永昕[1] 朱记法[1] 田俊[2] 王玉[2] 

机构地区：[1]华中科技大学协和医院心内科华中科技大学同济医学院血管病研究所 [2]华中科技大学同济医学院生物化学与分子生物学系

出　　处：《临床心血管病杂志》2004年第8期479-481,共3页Journal of Clinical Cardiology

基　　金：国家高技术研究发展计划 (86 3计划 )资助项目(No :2 0 0 2AA74 5 0 70)

摘　　要：目的 :探讨原代鼠骨骼肌成肌细胞异位表达重组人生长激素 (rhGH)的可行性。方法 :将目的基因人生长激素 (hGH)亚克隆至真核表达载体pcDNA3上 ,构建重组质粒pcDNA hGH。分别用酶切电泳及DNA测序的方法对重组质粒进行鉴定。用阳离子聚合物SofastTM介导的基因转移技术将重组质粒导入体外培养的原代大鼠骨骼肌成肌细胞。收集细胞培养上清液 ,用放射免疫分析 (RIA)的方法检测rhGH。结果 :酶切电泳和DNA测序显示hGHcDNA成功地插入到空载体中 ;rhGH表达持续 3周以上。结论 :该实验成功构建了能在真核细胞内表达的重组质粒pcDNA hGH ,为进一步研究以组织进行的基因治疗奠定了基础。Objective: To investigate the possibility of expression of recombinant human growth hormone (rhGH) in murine primary skeletal myoblasts. Method: Human growth hormone (hGH) cDNA was subcloned into expression vector pcDNA3 to construct recombinant pcDNA-hGH, identified by electrophoresis and sequencing analysis. Primary murine myoblasts were isolated, expanded in vitro, and transfected by cationic polymer Sofast TM reagent. RIA (radioimmuoassay) was used to detect the level of recombinant exogenetic protein delivered into medium. Result: HGH cDNA was correctly inserted into blank vector. Genetically modified myoblasts could secrete rhGH at least 3 weeks. Conclusion: Recombinant pcDNA-hGH has been constructed and expressed ectopic in murine myoblasts.

关 键 词：生长激素 成肌细胞 基因表达 

分 类 号：Q575.11[生物学—生物化学]