Immunoregulation

Immunoregulation

Recombinant Rabies Virus With a Multiple Cloning Site: A Platform Capable of Immunological Alterations

Document Type : Original Article

Authors
Seyed Hamidreza Naghi Mousavi 1
Alireza Gholami 2
Mojtaba Jafarinia 1
Mehdi Ajorloo 3
1 Department of Biology, Marvdasht Branch, Islamic Azad University, Marvdasht, Iran.
2 Viral Vaccines Production Unit, Research and Production Complex, Pasteur Institute of Iran, Tehran, Iran.
3 Hepatitis Research Center, Lorestan University of Medical Sciences, Khorramabad, Iran.
10.32598/Immunoregulation.4.2.5
Abstract
Background: Rabies is fatal encephalitis, i.e., preventable by appropriate vaccination. Reverse genetics has proved promising for manipulating the rabies virus immunological characteristics. The insertion or deletion of a gene from the rabies genome could render specific functions to the rabies virus. 
Materials and Methods: A multiple cloning site including 111 nucleotides long harboring 10 single-cut restriction sites have been designed. The designed fragment was cloned between the G and L genes of the rabies virus genome. The recombinant rabies virus was rescued, and its infectivity was confirmed in the BHK-21 cell line. The recombinant virus propagation was compared with the initial rabies virus strain. Statistical analysis was performed using GraphPad Prism.
Results: The cloning and localization of the multiple cloning site were verified by nucleotide sequencing. The recombinant virus properly propagated and rescued in the BHK-21 cell line. Comparing the recombinant virus with the initial rabies virus has shown that both viruses had similar functionality and propagation rate.
Conclusion: The recombinant virus obtained in the present study could facilitate further cloning experiments. Examples include constructing a marker virus, carrying green fluorescent protein to be used either in rabies immunity assays or tracking the virus infection in relevant tissues.
Keywords
Rabies virus
Reverse genetics
Virus engineering
Molecular cloning
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Immunoregulation
Volume 4, Issue 2
Winter 2021
Pages 101-108