【摘要】 目的：选择适当的载体，启动Ca²⁺依赖分泌型磷脂酶A2（SecretaryphospholipaseA2sPLA2）cDNA的表达，为进一步研究各种因素对该酶活性的影响奠定基础。方法：首先从sPLA2-pGEM7重组体（这一重组体不能在转染的真核细胞中表达）获得人sPLA2cDNA，克隆该片断进入中间载体BSSK（使aPLA2cDNA两端出现XbaI、Hind3酶切位点），选择antisensesPLA3-BSSK重组体进行扩增，双酶消化上述重组体及PRC／CMV，电洗脱回收aPLA2片断及线性化的载体PRC／CMV，定向连接sPLA2-PRC/CMV，重组体转染鼠巨噬细胞，筛选，扩增阳性克隆；提取RNA，Northem杂交法检测sPLA22mRNA。结果：应用定向克隆方法，成功地制备了sPLA2-PRC/CMV重组体。用这一重组质粒转染鼠巨噬细胞后，Northem杂交检测到了sPLA2－mRNA的表达。结论：SPLA₂－PRC／CMV是一有效的表达系统。更多还原

【Abstract】 Objective:Choose the effective vector for sPLA2cDNA and study the expression of sPLA2 in the vector.Method:we prepared the sPLA2cDNA from pGEM7; lagation of aPLA2-BSSk for getting different cohesive ends of sPLA2cDNA by Xbal HindIII: purifying the sPLA2-PRC/CMV;trasfacted sPLA2-PRC/CMV with mouse macrophage;isolatied RNA from macrophage cells,Northem blot;autoradiograph.Results:the orientation lagation of insert DNA is useful and essy to get recombinstion DNA;pGEM/is a expressing vector but the aPLA2 cDNA in it couldn’t be expressed .Conclusion:PRC/CMV is a useful expressing vector for aPLA2 cDNA更多还原