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目的构建人源性抗HBc单链抗体的真核表达载体并在细胞内表达。方法采用DNA重组技术将特异性人源性抗HBc单链抗体基因插入真核表达载体pEGFP-c1;转染HepG2细胞,经G418筛选细胞,荧光倒置显微镜观察细胞内抗HBc单链抗体与绿色荧光蛋白融合表达情况,并用ELISA法检测HBc单链抗体基因的细胞内表达。结果成功地构建了人源性抗HBc单链抗体的真核表达载体。转染HepG2细胞并筛选后,经荧光倒置显微镜观察,细胞内有绿色荧光蛋白表达;ELISA检测细胞内表达的单链抗体片段具有HBcAg结合活性。结论人源性抗HBc单链抗体的真核表达载体的构建并在细胞内成功表达,为胞内抗HBc单链抗体的进一步研究奠定了基础。
Objective To construct eukaryotic expression vector of human anti-HBc single chain antibody and express in cell. Methods The specific human anti-HBc single-chain antibody gene was inserted into the eukaryotic expression vector pEGFP-c1 by DNA recombination technique. The HepG2 cells were transfected into HepG2 cells and the cells were screened by G418. The anti-HBc scFv and green fluorescence Protein fusion expression, and HBc single chain antibody gene expression in the cell by ELISA. Results The eukaryotic expression vector of human anti-HBc single chain antibody was successfully constructed. After transfection of HepG2 cells and screening, the expression of green fluorescent protein in cells was observed by fluorescence inverted microscope. The single-chain antibody fragments expressed in the cells showed HBcAg binding activity by ELISA. Conclusion The construction of eukaryotic expression vector of human anti-HBc single chain antibody and its successful expression in the cell lay the foundation for the further study of intracellular anti-HBc single chain antibody.