Construction and Identification of Replicon of Contemporary Zika Virus Strain

Abstract

Zika virus (ZIKV) has caused significant outbreaks and epidemics in the Americas recently, and raised global concern due to its ability to cause microcephaly and other neurological complications. In this study, we constructed a subgenomic replicon of ZIKV
GZ01, which was a vector containing Renilla luciferase (RLUC) reporter gene, and was Japanese encephalitis pAJE70 as vector. On the basis of this, the RLUC reporter gene sequence was inserted into the coding structure protein gene position instead of the fragment encoding the structural protein (C, prM, E). The ZIKV GZ01 subgenomic replicon containing the RLUC reporter gene was constructed. After the plasmid was linearized, the replicon RNA was transcribed by sp6 promoter and transfected into BHK-21 cells. The replicons were identified by restriction endonuclease digestion with Not I/Kpn I and Hind III and the expression of non-structural protein NS1 was determined by indirect immunofluorescence assay. The luciferase activity was evaluated for viral replication. The results showed that the replicon RNA could express RLUC efficiently in BHK-21 cells,
and the RLUC activity increased linearly at 24–48 hours after transfection, which indicated that the replicon had good ability to express foreign genes. The construction of the subgenomic replicon of ZIKV GZ01 strain,which laid the foundation for the study of ZIKV replication and translation mechanism.