Virology
Construction of human adenovirus type 4 vector expressing enhanced green fluorescence protein
Xingui Tian, Yong Chen, Ye Fan, Zhichao Zhou, Shiying Chen, Wenkuan Liu, Rong Zhou
Published 2018-04-30
Cite as Chin J Microbiol Immunol, 2018, 38(4): 268-273. DOI: 10.3760/cma.j.issn.0254-5101.2018.04.005
Abstract
ObjectiveTo prepare human adenovirus type 4 (Ad4) vector expressing enhanced green fluorescence protein (EGFP).
MethodsThis study used a previously prepared plasmid pBRAd4 containing the whole genome DNA of Ad4-GZ01 strain. The Ad4 genome E3 region of pBRAd4 was deleted and replaced with the EGFP expression frame by conventional molecular cloning method. Then the recombinant plasmid was transfected into AD293 cells to rescue recombinant virus which was identified by sequencing, SDS-PAGE and ELISA. The purified virions were injected to mice and the induced immune responses were detected by ELISA and microneutralization test.
ResultsThe recombinant Ad4 vector rAd4EGFP expressing EGFP was obtained and could be recognized and neutralized by monoclonal antibody MN4b and antisera against Ad4. The Ad4-specific and EGFP-specific antibodies with high titers could be detected in mice immunized with rAd4EGFP.
ConclusionHuman Ad4 vector expressing EGFP was successfully obtained and could be used in research on vaccine development, drug evaluation and transgene vector.
Key words:
Human adenovirus type 4; Vector; Enhanced green fluorescence protein; Neutralization
Contributor Information
Xingui Tian
State Key Laboratory of Respiratory Disease, Guangzhou Institution of Respiratory Health, the First Affiliated Hospital of Guangzhou Medical University, Guangzhou Medical University, Guangzhou 510182, China
Yong Chen
Ye Fan
Zhichao Zhou
Shiying Chen
Wenkuan Liu
Rong Zhou
