关键词: 噬菌体, 抗体库, Cre-Loxp重组系统, 乙肝表面抗原

Abstract: Objective To construct an expression vector that can be applied to the construction of large Fab fragment phage display library.Methods BssH II restriction site in phagemid vector pDF was mutated to Bgl Ⅱ restriction sites by site-directed mutagenesis techniques, and a suicide gene SacB was inserted into both the heavy and light chain region, resulting in phagemid vector pDF-D-SacB. Heavy chain (VH) and light chain (VL) genes of anti-hepatitis B surface antigen (HBsAg) antibody were inserted into the phagemid vector pDF-D-SacB separately. The recombinant VH and VL phagemids were transformed into Trans1-Blue E. coli by electroporation respectively. By co-infecting the VH and VL phagemids into bacteria BSl365 at high multiplicity of infection, the recombinant phagemid containing both the VL and VH genes was obtained. After infecting trans1-Blue E. coli with the recombinant phage, the recombinant anti-HBsAg antibodies were assayed by ELISA. Results By inserting SacB gene into the light and heavy chain regions of pDF and modifying the cloning sites of the vector, a new phagemid vector pDF-D-SacB was constructed. The subsequent test showed that the constructed vector can express functional Fab phage antibodies, and can also allow the Cre-LoxP mediated recombination in bacteria cell. ConclusionA suicide gene containing phagemid vector pDF-D-SacB was applicable to constructing large phage display antibody library.

Key words: phage, antibody library, Cre-LoxP recombination system, HBsAg