Construction of recombinant viruses
Several gene fragments containing prM-E of ZIKV (accession: KX601168.1)
were designed (strategies shown in Fig. 1A). The gene fragment
“ZI-△-PA” was obtained by overlap PCR, in which the sequences from the
N-terminus to the C-terminus were prM-E of ZIKV (with deletion of the
stem-transmembrane (ST-TM) region), a linker and protein anchor 3 (PA3,
containing three lysin motifs). In ZI-JE-△-PA, the signal peptide of
ZI-△-PA was replaced with the signal peptide of Japanese encephalitis
virus (JEV) [14]. In ZI-GP-△-PA, the
signal peptide of ZI-△-PA was replaced with the gp67 signal peptide.
Signal peptide replacement was performed by PCR. All of the sequences
were optimized for insect cells and synthesized in pUC vector by Sangon
Biotech (Shanghai, China).
The above-mentioned target genes were cloned into the baculovirus
transfer vector pFastBac Dual under the pH (Not I + HindIII) and p10 (Sma I + Nhe I) promoters. Then, the vector
was transformed into E. coli DH10 Bac/AcMNPV competent cells, and
the recombinant baculovirus plasmid (bacmid) carrying two copies of the
target genes was obtained. The bacmid was transfected into Sf9 cells to
obtain recombinant baculovirus using Cellfectin II Reagent (Invitrogen
Co., Carlsbad, CA, USA) according to the manufacturer’s instructions.