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[目的]应用噬菌体展示技术构建人源性抗乙型肝炎(乙肝)表面抗原(HBsAg)单链抗体(ScFv)库。[方法]提取经免疫具有乙肝表面抗体的正常人淋巴细胞总RNA,采用反转录、触减PCR扩增人免疫球蛋白G(IgG)重链可变区基因(VH)和轻链可变区基因(VL),用Linker连接肽经重叠PCR将VH和VL基因连接成VH-Linker-VL形式的ScFv基因。将ScFv与pCANTAB-5E载体通过相同的酶切位点连接后,转化大肠杆菌TG1,经M13K07辅助噬菌体拯救,构建成全套人源性抗HBsAg ScFv库。[结果]成功构建了人源性抗HBsAg噬菌体ScFv库。[结论]人源性HBsAg噬菌体ScFv库的构建,为进一步筛选特异性高,具有治疗作用的乙肝抗体奠定基础。
[Objective] To construct human scFv library of human anti-hepatitis B surface antigen (HBsAg) using phage display technology. [Method] The total RNA of normal human lymphocytes immunized with hepatitis B surface antigen was extracted and the variable regions of human immunoglobulin G (IgG) heavy chain variable region (VH) and light chain variable The region genes (VL) were linked by overlapping PCR with Linker linker peptides to link the VH and VL genes to the ScFv gene in the VH-Linker-VL format. After connecting ScFv with pCANTAB-5E vector through the same restriction enzyme site, E.coli TG1 was transformed into E. coli TG1 and rescued by M13K07 helper phage to construct a full-length humanized anti-HBsAg ScFv library. [Result] The human anti-HBsAg phage ScFv library was successfully constructed. [Conclusion] The construction of humanized HBsAg phage ScFv library lays the foundation for further screening of specific and therapeutic hepatitis B antibodies.