关键词: 关键词：易错PCR, DNA-shuffling, 单链抗体库

Abstract: Abstract：Objective The high-storage capacity single-chain antibody (ScFv) library was constructed for selecting high-affinity antibodies using error-prone PCR and DNA-shuffling. Methods Collected 5ml blood from different age groups, gender and healthy people, mononuclear cells RNA were extracted, reverse transcription. VH and VL genes were amplified by PCR, and then VH and VL gene mutation amplified error-prone PCR. The full-length ScFv fragments were recombinated with the DNA-shuffling technology in vitro, and then transfer E.coli DH5α. The recombination rate was identified by colony PCR, antibody library diversity identified by restriction enzyme digestion. Results The single-chain antibody library was constructed by error-prone PCR and DNA-shuffling techniques. Analyzed by restriction enzyme digestion, the diversity of antibody library was good, the capacity was high storage. Conclusion The introduction of error-prone PCR and DNA-shuffling technology can help improve the genetic diversity, which will help build high-storage capacity of the single chain antibody library.

Key words: Keywords：Error-prone PCR, DNA-shuffling, ScFv library