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Integration of a hairpin RNA-encoding gene derived from Tobacco streak virus confers resistance in groundnut (Arachis hypogaea L.) against peanut stem necrosis disease

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Abstract

The feasibility of controlling peanut stem necrosis disease caused by Tobacco streak virus (TSV) in groundnut (Arachis hypogaea L.) was explored by expressing double-stranded RNA of the replicase (Rep) gene of TSV in groundnut through genetic engineering. A hairpin (hp) RNAi construct containing 535-bp sense and antisense TSV-Rep sequences flanking a 742-bp spacer sequence (Pdk intron) under the control of the constitutive Cauliflower mosaic virus 35S promoter was made in the binary vector pART27. This chimeric gene construct was then mobilized into Agrobacterium tumefaciens strain LBA4404 via triparental mating using pRK2013 as a helper. Cotyledon explants of groundnut cultivar TMV-7 were transformed with A. tumefaciens harboring the hpRNA cassette. The presence of the transgene in the transgenic plants was confirmed up to T3 generation by PCR amplification of the 535-bp fragment of TSV-Rep gene. The bioassay results indicated that necrotic lesions were observed on the leaves of the wild-type plants 7–9 days after inoculation with TSV and stem necrosis appeared 16–20 days after inoculation, whereas the transgenic plants did not develop symptoms until harvest. ELISA results indicated that the wild-type plants inoculated with TSV recorded the highest virus concentration as compared to the transgenic lines.

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Acknowledgments

We thank the CSIRO Plant Industry, Australia, for providing the vectors pHANNIBAL and pART27. We are grateful to Dr. S. Muthukrishnan, Department of Biochemistry & Molecular Biophysics, Kansas State University, Manhattan, Kansas, USA, for critical reading of the manuscript. This study was funded by the University Grants Commission, New Delhi, India (Grant No. 42-200/2013-SR).

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  1. Department of Plant Pathology, Centre for Plant Protection Studies, Tamil Nadu Agricultural University, Coimbatore, Tamil Nadu, 641 003, India

    Madam Gurivi Reddy, Chinnaiah Senthilraja, Rangasamy Adhithya, Vijayalakshmi Kothandaraman Satya, Ramalingam Rabindran & Rethinasamy Velazhahan

  2. Department of Plant Biotechnology, Centre for Plant Molecular Biology and Biotechnology, Tamil Nadu Agricultural University, Coimbatore, Tamil Nadu, 641 003, India

    Easwaran Kokiladevi & Durailagaraja Sudhakar

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  1. Madam Gurivi Reddy
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Correspondence to Rethinasamy Velazhahan.

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Reddy, M.G., Senthilraja, C., Adhithya, R. et al. Integration of a hairpin RNA-encoding gene derived from Tobacco streak virus confers resistance in groundnut (Arachis hypogaea L.) against peanut stem necrosis disease. J Plant Dis Prot 123, 205–214 (2016). https://doi.org/10.1007/s41348-016-0039-7

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