[Secreted expression of dengue virus type 2 envelope glycoprotein in eukaryotic cells].
Zhonghua Shi Yan He Lin Chuang Bing Du Xue Za Zhi
; 25(2): 85-8, 2011 Apr.
Article
en Zh
| MEDLINE
| ID: mdl-21863624
OBJECTIVE: To secreted express envelope glycoprotein (E) of dengue virus type 2 extracellularly. METHODS: The entire prM/E gene was amplified by RT-PCR. An optimized signal sequence gene from Japanese encephalits virus (JEV, SA14-14-2 strain) was introduced using fusion PCR. The impact of E protein transmembrane and cytoplasmatic domains was compared by amplifying prM and E with full length of E gene, with 20% truncation of the E gene at 3' terminus and one chimeric gene, which was generated by replacing the 3' terminal 20% region of E gene with the corresponding sequence of JEV (SA14-14-2 strain). The PCR segments were inserted into the NheI and NotI sites of pcDNA5/FRT vector or into the NheI and XhoI sites of pAcUW51-M. Then they were transfected into 293T cells or Sf9 cells respectively. The expression and secretion of E protein were detected by immunofluorescence assay (IFA) and Western Blot. RESULTS: After transected into 293T cells or Sf9 cells, all constructs expressed E protein intracellularly indentified by IFA while only two plasmids could secret detectable E protein into tissue culture using Western Blot analysis. CONCLUSION: Signal peptide as well as the transmembrane and cytoplasmatic domains is crucial for the secretion of dengue E protein.
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Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Expresión Génica
/
Proteínas del Envoltorio Viral
/
Dengue
/
Virus del Dengue
Límite:
Animals
/
Humans
Idioma:
Zh
Revista:
Zhonghua Shi Yan He Lin Chuang Bing Du Xue Za Zhi
Asunto de la revista:
VIROLOGIA
Año:
2011
Tipo del documento:
Article
País de afiliación:
China
Pais de publicación:
China