Construction and identification of non-immunized human phage display library
- VernacularTitle:人源性天然噬菌体展示文库的构建及鉴定
- Author:
Siji NIAN
;
Li HUANG
;
Xu WANG
;
Yuchuan WU
;
Qing YUAN
- Publication Type:Journal Article
- Keywords:
Single chain Fv fragment;
Phage antibody library;
Overlapping extension PCR
- From:
Chinese Journal of Immunology
1985;0(06):-
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To develop non-immunized human phage display library.Methods:The total RNA of lymphocyte cells from peripheral blood of healthy voluntee was isolated and cDNA was synthesized,and the genes of heavy variable chain (VH) and light variable chain (V? and V?) were amplified by direct PCR and half-nested PCR.By overlapping extension PCR,the genes of VH and VL (V? and V?) were linked.The linked genes of single chain Fv fragment (scFv) were ligated with the vector pCANTAB-5E and then cloned into TG1 for the scFv library construction.Results:By direct PCR and half-nested PCR,42 VH fragments,16 V? and 18 V? fragments were obtained.The size of linked scFv library genes was 750 bp and the volume of constructed scFv library was 1.35?108.The results of BstN Ⅰ analysis of scFv genes from the phage library showed that fingerprint map of the selected scFvs was different.Conclusion:The developed phage library is diversity and can be used for selecting humanized scFv.
