Abstract
Zika virus (ZIKV) has had detrimental effects on global public health in recent years. This is because the management of the disease has been limited, in part because its pathogenic mechanisms are not yet completely understood. Infectious clones are an important tool that utilize reverse genetics; these can be used to modify the ZIKV genomic RNA at the DNA level. A homologous recombination clone was used to construct pWSK29, a low copy plasmid that contained sequences for a T7 promoter, the whole genome of ZIKV ZKC2 strain, and a hepatitis delta virus ribozyme. High fidelity PCR was then used to amplify the T7 transcription template. The transcript was then transfected into susceptible cells via lipofection to recover the ZIKV ZKC2 strain. Finally, the virulence of rZKC2 was evaluated both in vitro and in vivo. The rZKC2 was successfully obtained and it showed the same virulence as its parent, the ZIKV ZKC2 strain (pZKC2), both in vitro and in vivo. The 3730 (NS2A-D62G) mutation site was identified as being important, since it had significant impacts on rZKC2 recovery. The 4015 (NS2A, A157V) mutation may reduce virus production by increasing the interferon type I response. In this study, one of the earliest strains of ZIKV that was imported into China was used for infectious clone construction and one possible site for antiviral medication development was discovered. The use of homologous recombination clones, of PCR products as templates for T7 transcription, and of lipofection for large RNA transfection could increase the efficiency of infectious clone construction. Our infectious clone provides an effective tool which can be used to explore the life cycle and medical treatment of ZIKV.
Highlights
Zika virus (ZIKV) is an important new mosquito-borne virus that is a member of the Flavivirus genus within the Flaviviridae family
The entire genome of the ZIKV ZKC2 strain was divided into four PCR fragments containing homologous sequences for homologous recombination
Fragment 4 was ligated to Hepatitis D ribozyme (HDVr) using overlapping PCR
Summary
Zika virus (ZIKV) is an important new mosquito-borne virus that is a member of the Flavivirus genus within the Flaviviridae family. Zika Virus Clone Construction on global public health (Heymann et al, 2016). Vaccines and antiviral medications have not yet been developed for ZIKV and its pathogenic mechanisms are not yet fully understood. Reverse genetic systems can recover viruses containing specific mutations. They are important when studying RNA viruses, as they can modify them at the DNA level, providing a powerful tool for studying the pathogen’s life cycle, and pathogenic and virus–host cell interaction mechanisms. Constructing infectious clones of ZIKV is an indispensable experimental technique that can be used to help improve our understanding of pathogen biology and investigate related treatment methods
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
