EvaGreen-based Multiplex Real-time PCR Assay for Rapid Differentiation of Wild-Type and Glycoprotein E-Deleted Bovine Herpesvirus-1 Strains |
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| Authors: | Sachin S Pawar Chetan D Meshram Niraj K Singh Mohini Saini B P Mishra Praveen K Gupta |
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| Institution: | 1. Division of Veterinary Biotechnology, Indian Veterinary Research Institute, Izatnagar, India;2. School of Animal Biotechnology, Guru Angad Dev Veterinary and Animal Sciences University, Ludhiana, India;3. Division of Biochemistry, Indian Veterinary Research Institute, Izatnagar, India |
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| Abstract: | Bovine herpesvirus-1 (BoHV-1) is an important viral pathogen causing significant economic losses to the cattle industry. Glycoprotein E-deleted marker vaccines form the basis for BoHV-1 control programs widely, wherein detection and differentiation of wild-type and gE-deleted vaccine strains is of crucial importance for proper disease management. In the present study, we report an EvaGreen-based multiplex real-time polymerase chain reaction (EGRT-PCR) assay for rapid differentiation of wild-type and glycoprotein E-deleted strains of BoHV-1. The EGRT-PCR assay could simultaneously detect two viral genes (glycoprotein B and E) and an internal positive control gene (bovine growth hormone- bGH), in a single-tube reaction. The analytical sensitivity of the EGRT-PCR assay was as little as 10 copies of the BoHV-1 DNA per reaction. The modified real-time PCR assay could successfully differentiate wild-type and gE-deleted BoHV-1 strains based on gene specific melting temperatures (Tm) peaks. Our results have shown that the EGRT-PCR developed in this study might prove to be a promising tool in disease management by enabling rapid differentiation of wild-type and gE-deleted strains of BoHV-1. |
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| Keywords: | Bovine herpesvirus-1 differentiation EvaGreen wild-type |
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