[Secreted expression of dengue virus type I envelope glycoprotein in 293T cells].
OBJECTIVE
To expression prM/E gene of dengue virus type I in mammalia cells.


METHODS
The full-length prM/E gene of dengue virus type I strain GZ01/95 was amplified by RT-PCR, the signal peptide preceding the prM gene was added or the carboxyl-terminal 20% of DEN-1 E was replaced with the corresponding JE sequence in the meanwhile, and three of the constructions were cloned into the pcDNA5/FRT.Then they were transfected into 293T cells by lipofectamine respectively. The expression of recombinant proteins were identified by indirect immuno-fluorescence assay(IFA) as well as Western blot.


RESULTS
In the cytoplasm of 293T cells transfected with all the recombinant plasmids DNA, the expressed products for gene of dengue virus type I were confirmed by IFA. The secreted expression products for gene of dengue virus type I specific protein bands were confirmed by Western blot only existing in the cell supernatants transfected with the modified recombinant plasmids DNA.


CONCLUSIONS
The prM/E protein of dengue virus type 1 were expressed in 293T cells transfected with all the three recombinant plasmids DNA. The prM/E protein was obtained secretion after transfecting the modified recombinant plasmids adding a signal peptide preceding the prM gene or replacing the carboxyl-terminal 20% of E with the corresponding JE sequence.
Miao
Fang
F
National Institute for Viral Disease Control and Prevention, Chinese Center for Disease Control and Prevention, State Key Laboratory for Molecular Virology & Genetic Engineering, Beijing 100052, China.
Li
Chuan
C
Zhang
Shuo
S
Wang
Xiao-fang
XF
Li
Jian-dong
JD
Zhang
Quan-fu
QF
Liu
Qin-zhi
QZ
Wei
Yan
Y
Hang
Xiao-tong
XT
Liang
Mi-fang
MF
Li
De-xin
DX
D002460:Cell Line; D003715:Dengue / Q000821:virology*; D003716:Dengue Virus / Q000235:genetics* / Q000378:metabolism; D015870:Gene Expression; D006023:Glycoproteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D021381:Protein Transport; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D014759:Viral Envelope Proteins / Q000235:genetics* / Q000378:metabolism*
C084188:E protein, Dengue virus type 3; D006023:Glycoproteins; D011993:Recombinant Fusion Proteins; D014759:Viral Envelope Proteins; C023587:protein M (glycoprotein)