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RNAi-based transgene conferred extreme resistance to the geminivirus causing apical leaf curl disease in potato

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Abstract

Potato apical leaf curl disease is an emerging geminiviral disease in tropics and subtropics. It was reported for the first time in the year 1999 in northern plains of India but quickly spread to almost all potato growing regions of the country largely due to prevalence of warmer weather during early crop growth, thereby favoring whitefly vector. The problem of apical leaf curl disease in India became more severe due to lack of seed indexing for this virus in conventional seed production scheme. Although it accounts for major yield loss, there is no conventional source of resistance available in potato against Tomato Leaf Curl New Delhi Virus-Potato (ToLCNDV-Potato) that causes this disease in potato. In the present study, we have investigated the potential use of RNAi for obtaining resistance against this DNA virus in potato. The replication-associated protein gene (AC1) of the virus was used to obtain pathogen-derived resistance. The AC1 gene was PCR amplified from field-infected potato leaves, cloned and sequenced (JN393309). It showed 93% sequence similarity with the AC1 gene of Tomato Leaf Curl Virus-New Delhi (TOLCV-NDe; DQ169056) virus. Transgenic plants encoding the AC1 gene in three different orientations, viz. sense, antisense and hairpin loop, were raised. Transgenic lines when challenge inoculated with ToLCNDV-Potato showed different levels of resistance for all three constructs. Transgene integration and copy number in selected transgenic lines were determined by qPCR and further confirmed by Southern blot analysis. Though a reduction in viral titer was observed in transgenic lines encoding either antisense or hairpin loop constructs of AC1 gene, the latter transgenics showed most significant results as shown by reduction in the level of symptom expression in glasshouse screening as well as real-time data of in vivo virus concentration. In fact, we obtained a few totally asymptomatic transgenic lines with hairpin loop strategy.

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Acknowledgements

The authors acknowledge the financial grant received from Indian Council of Agricultural Research, New Delhi, India. We are also thankful to Drs. S. K. Pandey and B. P. Singh, Former Directors, ICAR-CPRI, Shimla, for providing laboratory facilities for conducting this study.

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Authors and Affiliations

  1. ICAR-Central Potato Research Institute, Shimla, Himachal Pradesh, 171001, India

    Garima Tomar, S. K. Chakrabarti, Nitya Nanda Sharma, A. Jeevalatha, S. Sundaresha & Kanika Vyas

  2. Himachal Pradesh University, Summer Hill, Shimla, 171005, India

    Wamik Azmi

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  1. Garima Tomar
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  2. S. K. Chakrabarti
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  3. Nitya Nanda Sharma
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  5. S. Sundaresha
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  6. Kanika Vyas
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Correspondence to S. Sundaresha.

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Garima Tomar, S. K. Chakrabarti, Nitya Nand Sharma, A. Jeevalatha, S. Sundaresha, Kanika Vyas, and Wamik Azmi declare that they have no conflict of interest.

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This article does not contain any studies with human participants or animals performed by any of the authors.

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Tomar, G., Chakrabarti, S.K., Sharma, N.N. et al. RNAi-based transgene conferred extreme resistance to the geminivirus causing apical leaf curl disease in potato. Plant Biotechnol Rep 12, 195–205 (2018). https://doi.org/10.1007/s11816-018-0485-8

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