Secreted expression of dengue virus type 2 envelope glycoprotein in Eukaryotic cells
10.3760/cma.j.issn.1003-9279.2011.02.002
- VernacularTitle:登革2型病毒ZS01/01株E蛋白在真核细胞中的分泌表达研究
- Author:
Shuo ZHANG
1
;
Wen GU
;
Chuan LI
;
Fang MIAO
;
Peng LU
;
Jing QU
;
Yan WEI
;
Quan-Fu ZHANG
;
Qin-Zhi LIU
;
Jian-Dong LI
;
Mi-Fang LIANG
;
De-Xin LI
Author Information
1. 中国疾病预防控制中心病毒病预防控制所
- Keywords:
Dengue virus;
Viral structural protein;
Expresion sequence tags/secretion
- From:
Chinese Journal of Experimental and Clinical Virology
2011;25(2):85-88
- CountryChina
- Language:Chinese
-
Abstract:
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 Notl sites of pcDNA5/FRT vector or into the Nhel and Xhol sites of pAcUW51-M. Then they were transfected into 293T cells or St9 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 St9 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 transmemhrane and cytoplasmatic domains is crucial for the secretion of dengue E protein.
