Generation of infectious clone of bovine adenovirus type I expressing a visible marker gene

Abstract

Background and objective: Bovine adenovirus type 3 (BAdV3) has been widely used as a vector for vaccine research and development, whereas BAdV1 biology and BAdV1-based vectored vaccine have been less frequently reported. We aimed to construct an infectious BAdV1 clone and explore the functions of BAdV1 genes. Methods: First, the infectious clone of pUCBAdV1 containing the full-length BAdV1 DNA and the recombinant plasmid pUCBAV1-EYFP expressing the marker gene EYFP were constructed. Then, the recombinant viruses BAdV101 and rBAdV1-EYFP were rescued. The stability of the exogenous EYFP gene was analyzed by continuous passage, PCR, and western blotting. Finally, the virus neutralization titer of the rescued viruses was evaluated. Results: The infectious clones of pUCBAdV1 and pUCBAV1-EYFP were constructed and the recombinant viruses BAdV101 and rBAdV1-EYFP were rescued successfully. Moreover, the results showed that the EYFP gene could be expressed continuously. In addition, the replication of rBAdV1-EYFP was less efficient than that of the wild-type virus wtBAdV1 in vitro, while the efficacy of BAdV101 replication was almost the same as that of wtBAdV1. Furthermore, the neutralization test showed that the neutralization titer of rBAdV1-EYFP was consistent with that of wtBAdV1. Conclusion: To our knowledge, the infectious genome of pUCBAV1-EYFP expressing a visible marker gene EYFP was constructed for the first time, and the finding forms a basis for the development of BAdV1-based efficient vectored vaccine.