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Viruses 2019, 11(2), 115; https://doi.org/10.3390/v11020115

New Method for Differentiation of Granuloviruses (Betabaculoviruses) Based on Real-Time Polymerase Chain Reaction (Real-Time PCR)

1
Laboratory of Recombinant Vaccines, Intercollegiate Faculty of Biotechnology of University of Gdansk and Medical University of Gdansk, University of Gdansk, 80-307 Gdansk, Poland
2
Department of Biochemistry and Microbiology, Rhodes University, P.O. Box 94, Grahamstown 6140, South Africa
3
Centre for Biological Control, Department of Zoology and Entomology, Rhodes University, P.O. Box 94, Grahamstown 6140, South Africa
4
Embrapa Recursos Genéticos e Biotecnologia, Parque Estacao Biológica, Brasilia 70770-900, Brazil
5
Citrus Research International (CRI), P.O. Box 5095, Walmer 6065, Port Elizabeth, South Africa
*
Author to whom correspondence should be addressed.
Received: 23 December 2018 / Revised: 24 January 2019 / Accepted: 24 January 2019 / Published: 29 January 2019
(This article belongs to the Special Issue Insect Viruses and Pest Management)
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Abstract

Baculoviridae is a highly diverse family of rod-shaped viruses with double-stranded DNA. To date, almost 100 species have had their complete genomic sequences deposited in the GenBank database, a quarter of which comprises granuloviruses (GVs). Many of the genomes are sequenced using next-generation sequencing, which is currently considered the best method for characterizing new species, but it is time-consuming and expensive. Baculoviruses form a safe alternative to overused chemical pesticides and therefore there is a constant need for identifying new species that can be active components of novel biological insecticides. In this study, we have described a fast and reliable method for the detection of new and differentiation of previously analyzed granulovirus species based on a real-time polymerase chain reaction (PCR) technique with melting point curve analysis. The sequences of highly conserved baculovirus genes, such as granulin and late expression factors 8 and 9 (lef-8 and lef-9), derived from GVs available to date have been analyzed and used for degenerate primer design. The developed method was tested on a representative group of eight betabaculoviruses with comparisons of melting temperatures to allow for quick and preliminary granulovirus detection. The proposed real-time PCR procedure may be a very useful tool as an easily accessible screening method in a majority of laboratories. View Full-Text
Keywords: betabaculovirus; detection; real-time PCR; lef-9; lef-8; granulin betabaculovirus; detection; real-time PCR; lef-9; lef-8; granulin
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Krejmer-Rabalska, M.; Rabalski, L.; Jukes, M.D.; Lobo de Souza, M.; Moore, S.D.; Szewczyk, B. New Method for Differentiation of Granuloviruses (Betabaculoviruses) Based on Real-Time Polymerase Chain Reaction (Real-Time PCR). Viruses 2019, 11, 115.

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