Identification of the optimal insertion site for expression of a foreign gene in an infectious hematopoietic necrosis virus vector
Infectious hematopoietic necrosis virus (IHNV) was developed as a vector to aid the construction of vaccines against viral diseases such as viral hemorrhagic septicemia virus, spring viremia of carp virus, and influenza virus H1N1. However, the optimal site for foreign gene expression in the IHNV vector has not been determined. In the present study, five recombinant viruses with the green fluorescence protein (GFP) gene inserted into different genomic junction regions of the IHNV genomic sequence were generated using reverse genetics technology. Viral growth was severely delayed when the GFP gene was inserted into the intergenic region between the N and P genes. Real-time fluorescence quantitative PCR assays showed that the closer the GFP gene was inserted towards the 3ʹ end, the higher the GFP mRNA levels. Measurement of the GFP fluorescence intensity, which is the most direct method to determine the GFP protein expression level, showed that the highest GFP protein level was obtained when the gene was inserted into the intergenic region between the P and M genes. The results of this study suggest that the P and M gene junction region is the optimal site within the IHNV vector to express foreign genes, providing valuable information for the future development of live vector vaccines.
This study was supported by the Central Public-Interest Scientific Institution Basal Research Fund, CAFS (grant number HSY201804M and 2019ZD0704), the Natural Science Foundation of Heilongjiang Province (grant number QC2018039), the National Natural Science Foundation of China Grant (grant number 31802344), and the Heilongjiang Postdoctoral Program (LBH-Z18275).
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Conflict of interest
The authors declare that they have no conflict of interest related to this work.
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