Change in the Basic Structure of the Rabies Virus Glycoprotein by Reverse Genetics

Rabies is a deadly zoonotic disease that is caused by the rabies virus. The virus can infect and disrupt the central nervous system of a rabid patient. The rabies virus is a neurotropic single stranded RNA virus. Glycoprotein (G) is the most important protein that binds to the cellular receptors and also induces an immune response against the virus in the host. Using reverse genetics technology, the glycoprotein gene could be modified and a virus with higher immunogenicity or lower pathogenicity.

In this study, we designed a mutation in the sequence of glycoprotein gene using a software, on the main antigenic site II of the Pasteur virus strain at the position of 42-34 amino acids. Agene fragment in the cloning vector containing the rabies virus genome was replaced by the synthesized construct containing the altered gene by two restricted enzymes, and then cloned. The T7-BHK cell under the T7 phage promoter control was transfected to express the glycoprotein gene, along with the construct and vectors expressing the N, P, and L genes of the rabies virus as well as the full genome. After expressing and confirming viral genes, it was cultured and amplified in BSR cell.

after cloning and expression of the recombinant virus in the target cell, the vector containing the mutated gene led to the rescue of the recombinant virus. The recombinant virus cultured and propagated in the BSR cells, then the genome was extracted and finally confirmed by sequencing.

The rescued recombinant virus can be used for research studies or in the vaccines manufacturing, provide that the antigenicity is maintained or increased.