人甘露糖结合凝集素在小鼠肝脏细胞内表达

目的：在小鼠肝脏组织中表达人甘露糖结合凝集素（mannose-binding lectin，MBL）。方法：采用PCR方法从pGEMT-MBL质粒中扩增人MBLcDNA序列，克隆人pUC19载体中，限制性酶切鉴定及测序正确后，将MBLcDNA克隆人真核表达载体pcDNA3质粒中，构建成pcDNA3-MBL质粒，经尾静脉大容量快速注射pcDNA3-MBL质粒20μg，8h后处死小鼠，Western Blot方法检测小鼠肝脏组织和血清中人MBL，免疫组织化学方法检测小鼠肝脏组织中人MBL的表达情况。结果：成功扩增出人MBLcDNA序列并构建克隆载体pUC19-MBL，测序结果正确，成功构建pcDNA3-MBL真核表达载体。小鼠尾静脉注射pcDNA3-MBL质粒后8h，免疫组化检测证实肝细胞中有人MBL表达，Western blot方法检测发现血清和肝脏组织中出现人MBL蛋白。结论：真核表达载体pcDNA3-MBL裸质粒DNA静脉注射可以在小鼠肝脏中大量表达人MBL，并分泌人血，为临床纠正先天性低MBL患者的易感染状态提供了新的思路。

Expression of hmnan mannose-binding lectin in mice liver

Objective:To amplify and clone the human mannose-binding lectin(MBL) gene and to express its recombinant protein in mouse liver tissue.Methods:After the human MBL cDNA was amplified by PCR and identified by sequencing and restriction mapping,it was inserted into eukaryotic expression vector pcDNA3.The recombinant plasmid pcDNA3-MBL was injected into mice in large quantity and large volume in a short time through tail vein.These mice were sacrificed 8 h after the injection.MBL were examined in serum and hepatic tissue with Western blot and immunohistochemistry.Results:After the human MBL gene was amplified and sequenced correctly,it was successfully inserted in the eukaryotic expression vector pcDNA3.Eight hours after the pcDNA3-MBL plasmid were injected into mice through tail veins,human MBL could be found both in serum and hepatic tissue with Western blot and immunohistochemistry examination.Conclusion:Systemic injection of pcDNA-MBL can result in human MBL protein expression in mice liver and secretion into blood.This result may provide a new idea to treat the congenital MBL insufficient patients,who have the predisposition of infectious diseases.