Construction and expression of HSV-2gD-Hsp70 fusion protein gene.
- Author:
Jian-Yong FAN
1
;
Hui-Lan YANG
;
Ying WANG
;
Lei GUAN
Author Information
1. Department of Dermatology, Guangzhou General Hospital, Guangzhou 510010, China. fanjianyong@hotmail.com
- Publication Type:Journal Article
- MeSH:
Animals;
Blotting, Western;
Cell Proliferation;
Gene Expression;
HSP70 Heat-Shock Proteins;
biosynthesis;
genetics;
immunology;
isolation & purification;
Immunoglobulin G;
blood;
immunology;
Interferon-gamma;
blood;
Mice;
Plasmids;
genetics;
Recombinant Fusion Proteins;
biosynthesis;
genetics;
immunology;
isolation & purification;
Spleen;
cytology;
immunology;
Viral Envelope Proteins;
biosynthesis;
genetics;
immunology;
isolation & purification
- From:
Chinese Journal of Biotechnology
2006;22(6):914-918
- CountryChina
- Language:Chinese
-
Abstract:
To construct and express Hsp70-HSV2gD fusion protein. Genes of Hsp70 and HSV-2gD were subcloned into vectors pGEX-4T-1 respectively. After confirmed by DNA sequence analysis, the recombinant plasmids pGEX-4T-HSP-gD was transformed into E. coli DH5alpha and induced to express with IPTG. The expressed protein was characterized by SDS-PAGE and Western blot after purified. BALB/c mice were immunized with fusion proteins respectively via intra-m uscular injection. The proliferation of spleen lymphocytes, the level of y-IFN in culture and anti-HSV-2gD IgG antibody in serum was detected was detected. The expressed protein was analyzed by SDS-PAGE after induced with IPTG, which showed a new band with an apparent molecular mass corresponding to the predicted size (118 kD). Western Blotting analysis demonstrates that the purified Hsp70-HSV2gD fusion protein had specific binding activity. The stimulation indexes of spleen lymphocytes, the level of gamma-IFN in culture and anti-HSV-2gD IgG antibody in serum of GST-Hsp70-gD group was obviously higher than that of other groups (P < 0.05 respectively). The successful expression of the Hsp70-HSV2gD fusion protein, which can induce immune responses, laid a solid foundation for its further research.
