摘要
目的 从人正常肺组织中提取羧肽酶 A基因后进行了定点突变 ,并将突变型人羧肽酶 A基因与抗精浆蛋白单链抗体基因进行融合 .方法 应用重叠延伸法将突变型人羧肽酶 A与抗精浆蛋白单链抗体基因进行融合 ,并在大肠杆菌中进行表达 .结果 序列分析表明 :我们得到大约为 1 980 bp的基因 ,其 5 端为抗精浆蛋白单链抗体基因 ,3 端为人羧肽酶 A突变体基因 ,抗精浆蛋白单链抗体和突变型人羧肽酶 A基因成功地融合为一条基因 ,应用 p GEX- 4 T- 1表达载体原核表达得到一 Mr约为 970 0 0的特异蛋白 .结论 抗精浆蛋白单链抗体和突变型人羧肽酶
AIM To make point mutation hCPA gene and construct, express a fusion gene of γ seminoprotein ScFv/ carboxypeptidase A mutant gene. METHODS We constructed a fusion gene of γ seminoprotein ScFv/carboxypeptidase A mutant gene by over lap extension PCR. The fusion gene was expressed in E.coli . RESULTS Sequencing indicated that the product of over lap extension PCR consisted of 1980 bp, 5 end of the fusion gene was γ seminoprotein ScFv and 3 end was hCPA gene. After induction for 3 hours in E.coli , a new anticipatory protein band appeared on SDS PAGE gel. CONCLUSION Fusion gene of γ seminoprotein ScFv/carboxypeptidase A mutant gene is constructed and expressed successfully in E.coli . It might provide foundation for application of hCPA in ADEPT.
出处
《第四军医大学学报》
北大核心
2002年第4期304-307,共4页
Journal of the Fourth Military Medical University
基金
高等学校骨干教师资助计划资助(2000-65-66)
关键词
单链抗体
羧肽酶类
突变
基因表达
前列腺癌
single chain antibody
carboxypeptidases, mutation
gene expression
