Construction of gene library of human-originated single-chain variable fragment antibody associated with lung adenocarcinoma
- VernacularTitle:人源肺腺癌单链抗体基因文库的构建
- Author:
Hua PANG
- Publication Type:Journal Article
- Keywords:
Phage antibody library;
Single-chain variable fragment;
Lung adenocarcinoma;
Construction
- From:
Journal of Chongqing Medical University
2007;0(11):-
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To construct human single-chain variable fragment (scFv) antibodies gene library associated with lung adenocarcinoma. Methods:Total RNA was extracted from the lymphatic tissue near the lung adenocarcinoma.VH and VL fragments were amplified with RT-PCR,the purified VH and VL fragments were used to produce scFv fragment by splicing-overlap-extension PCR. The scFv gene was cloned in pCANTAB-5E Phasmid,then transferred to Ecoli TG1.The insertion ratio of antibody gene was identified with RT-PCR,the positive enzyme-cutting products were identified with 1.5% agarose gel electrophoresis. Results:Clear 28 S and 18 S strips were found in total RNA;The size of VH is about 370 bp,VL is about 350 bp.The size of scFv is about 750 bp.After transformation into Ecoli TG1,2.2?107 CFU/?g ampicillin resistant bacteria colonies grow after overnight culture,and the positive insert ratio was 83.3%. Conclusion:Human single-chain variable fragment(scFv) antibodies gene associated with lung adenocarcinoma was constructed successfully. It could provide bases to further screen the antibody library construction.
