Construction of herpes simplex virus type I glycoprotein D DNA vaccine and its preliminary study / 中国医学科学院学报
Acta Academiae Medicinae Sinicae
;
(6): 67-70, 2002.
Artículo
en Chino
| WPRIM
| ID: wpr-280963
ABSTRACT
<p><b>OBJECTIVE</b>The goal of this study was to construct a eukaryotic expression plasmid containing the gene encoding herpes simplex virus type I glycoprotein D (HSV-1, gD) and evaluate its utility for DNA immunization in mice.</p><p><b>METHODS</b>The gD gene was amplified from viral DNA using PCR with EcoR I and BamH I restriction sites encoded on 5' and 3' ends, respectively. The PCR fragment was inserted into the transfer vector pGEM-T Easy. gD was then cut from this vector and inserted into the EcoR I and BamH I sites in the pcDNA3.1 at the multiple cloning sites (MCS). The recombinant plasmid, pcDNA3.1-gD1, was transfected into COS-7 cells using Lipofectamine according to the manufacture's instructions. The expression of the glycoprotein D was analyzed by immunoblotting of the cell lysates. 4-6 weeks old BALB/C mice were given two injections at tibia anterialis muscle, each containing 100 micrograms of plasmid DNA, on days 0 and 15. pcDNA3.1 was used as negative control. Blood samples were taken from all mice at weeks 0, 2, 4, and 6 after the first inoculation. Standard indirect ELISA was employed to evaluate the levels of specific total Ig in serum.</p><p><b>RESULTS</b>The recombinant plasmid was confirmed with restriction digestion and sequencing to contain target gene segment and expressed in COS-7 cells in vitro shown by Western blotting. The pcDNA3.1-gD1 immunized group induced specific antibody response as compared to the negative control, and the titer was about 12000.</p><p><b>CONCLUSIONS</b>The recombinant plasmid pcDNA3.1-gD1 is potential to be used as a candidate vaccine, for the treatment of HSV-1 infection.</p>
Texto completo:
Disponible
Índice:
WPRIM (Pacífico Occidental)
Asunto principal:
Proteínas del Envoltorio Viral
/
Células COS
/
Vacunas de ADN
/
Alergia e Inmunología
/
Escherichia coli
/
Vectores Genéticos
/
Genética
/
Metabolismo
/
Ratones Endogámicos BALB C
Límite:
Animales
/
Humanos
Idioma:
Chino
Revista:
Acta Academiae Medicinae Sinicae
Año:
2002
Tipo del documento:
Artículo
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