Identification and functional analysis of the GTPV bidirectional promoter region
The goat pox chick embryo-attenuated virus (GTPV) has been developed as an effective vaccine that can elicit protective immune responses. It possesses a large genome and a robust ability to express exogenous genes. Thus, this virus is an ideal vector for recombinant live vaccines for infectious diseases in ruminant animals. In this study, we identified a novel bidirectional promoter region of GTPV through screening named PbVV(±). PbVV(±) is located between ETF-l and VITF-3, which are transcribed in opposite directions. A new recombinant goat pox virus (rGTPV) was constructed, in which duplicate PbVV(+) was used as a promoter element to enhance Brucella OMP31 expression, and duplicate PbVV(−) was used as a promoter element to regulate enhanced green fluorescent protein (EGFP) at the same time as the selection marker. PbVV(−) promoter activity was compared to that of the P7.5 promoter of vaccinia virus, as measured by EGFP expression; the fluorescence intensity of EGFP expressed in cells was confirmed by fluorescence microscopy and flow cytometry. PbVV(+) promoter activity was measured by Brucella OMP31 expression. Interaction with the anti-Brucella-OMP31 monoclonal antibody was confirmed by western blotting, and OMP31 mRNA expression was assessed by qRT-PCR. The results of this study will be useful for the further study of effective multivalent vaccines based on rGTPV. This study also provides a theoretical basis for overcoming the problem of low expression of exogenous genes.
KeywordsGoat pox virus Bidirectional promoter Vector Omp31
This study was supported by the International Science and Technology Cooperation Project of China (2013DFR30970), the National Science and Technology Major Project (2013BAI05B05), the National Natural Science Foundation of China (U1303283, 31360610, 31602080), and the University Key Research Project of Henan Province (16A230013).
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