Development of a recombinase polymerase amplification assay for the diagnosis of banana bunchy top virus in different banana cultivars
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Recombinase polymerase amplification (RPA) is a rapid, isothermal amplification method with high specificity and sensitivity. In this study, an assay was developed and evaluated for the detection of banana bunchy top virus (BBTV) in infected banana plants. Three oligonucleotide primer pairs were designed from the replicase initiator protein gene sequences of BBTV to function both in RPA as well as in polymerase chain reaction (PCR). A total of 133 symptomatic as well as asymptomatic banana leaf samples from various cultivars were collected from the different regions of India and evaluated for BBTV infection using the RPA assay. BBTV was efficiently detected using crude leaf sap in RPA and the results obtained were consistent with PCR-based detection using purified DNA as template. To our knowledge, this is the first report of reliable diagnosis of BBTV infection by RPA using crude leaf sap as a template.
KeywordsPolymerase Chain Reaction Conventional Polymerase Chain Reaction Recombinase Polymerase Amplification Crude Leaf Extract Banana Leaf
This study was supported by the Department of Biotechnology under the Referral Centre of “National Certification System of Tissue Culture Raised Plants” (NCS-TCP). The authors wish to thank the Head of the Division of Plant Pathology and Director of the Indian Agricultural Research Institute, for providing the necessary lab facilities. We are also grateful to Dr. Akhilesh Mishra of Jain Irrigation Systems Ltd, Jalgaon, and Dr. Vimi Louis, Associate Professor at Kerala Agricultural University, for help with sample collection.
Compliance with ethical standards
This study was funded by the Department of Biotechnology, Government of India, under the National Certification System for Tissue Culture Raised Plants project.
Conflict of interest
The authors declare that they have no competing interests.
This article does not contain any studies with human participants or animals performed by any of the authors
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