The construction and expression of superantigen SEA and antimelanoma ScFv fusion gene.
- Author:
Jing SUN
1
;
An-Guo LÜ
;
Wen-Fang WU
;
Xiang-Yang BAI
;
Xiu-Bao REN
;
Hong LIU
Author Information
1. Shenyang Institute of Applied Ecology of Chinese Academy of Science, Shenyang 110015, China.
- Publication Type:Journal Article
- MeSH:
Cell Line, Tumor;
Cell Survival;
drug effects;
Cells, Cultured;
Electrophoresis, Polyacrylamide Gel;
Enterotoxins;
genetics;
metabolism;
Escherichia coli;
genetics;
metabolism;
Humans;
Inclusion Bodies;
genetics;
metabolism;
Melanoma;
drug therapy;
immunology;
Recombinant Fusion Proteins;
genetics;
metabolism;
pharmacology;
therapeutic use;
Single-Chain Antibodies;
genetics;
metabolism
- From:
Chinese Journal of Biotechnology
2003;19(6):750-753
- CountryChina
- Language:Chinese
-
Abstract:
Two strategies, direct ligation after enzyme digestion and over-lap PCR technology, were adopted to construct a fusion gene which was composed of the antimelanoma single chain antibody gene and the staphylococcal enterotoxin A gene without N-terminal signal sequence. The fusion gene was subcloned into pET28-a vector and transformed into E. coli BL21(DE3). Ni-NTA system was selected to separate and purify the expresstd products. The inhibition ratio of the fusion protein was tested by MTT method. It is shown that the 6His-ScFv-SEA fusion protein can be expressed stably in E. coli BL21 (DE3). The quantity of the fusion protein was shown up to 30% of the total protein of the bacteria and mainly in inclusion body. By activation the effective cells, the fution protein can inhibit the melanoma cell whith expressed corresponding antigen.
