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Targeting genes involved in nucleopolyhedrovirus DNA multiplication through RNA interference technology to induce resistance against the virus in silkworms

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Abstract

RNA interference (RNAi) has become an efficient tool for inducing resistance to viruses in many organisms. In this study, Escherichia coli cells were engineered to produce stable double-stranded RNA (dsRNA) against the nucleopolyhedrosis virus to elicit RNAi in silkworms. The immediate-early-1 (ie-1) and late expression factor-1 (lef-1) genes of the Bombyx mori nucleopolyhedrovirus (BmNPV) involved in viral DNA multiplication were cloned in the plasmid L4440 under the influence of the double T7 promoter and transformed to E. coli HT115 DE3 host cells. On induction with isopropyl β-d-thiogalactopyranoside, these cells efficiently produced dsRNA of the cloned genes. The B. mori larvae were fed with 50 µL of E. coli cells expressing ie-1 and lef-1 dsRNAs (each approximately 25 µg) to elicit RNAi. The semi-quantitative and quantitative PCR analysis of RNA from the midgut of the dsRNA-fed larvae revealed a significant reduction in the expression of the target genes involved in BmNPV multiplication, which restricted virus copy numbers to 100 compared with 1.9 × 105 in the infected controls. Furthermore, the dsRNA-fed infected larvae showed > 50% increased survivability compared with the infected controls. The study revealed the successful use of bacteria as vectors for efficiently delivering dsRNA to elicit RNAi against BmNPV in silkworms.

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Acknowledgements

This Research was funded by Central Silk Board, Bengaluru and Technical Collaboration was provided by Professor M.V. Rajam, Department of Genetics, UDSC, New Delhi. The authors would like to acknowledge Dr. Geetha Murthy, Scientist, CSGRC, Hosur and Editingindia for meticulously editing the Manuscript.

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  1. Sahar Ismail, and K. S. Tulsi Naik contributed equally to the work

Authors and Affiliations

  1. Seri Biotech Research Laboratory, Central Silk Board, Kodathi, Bengaluru, Karnataka, 560035, India

    Sahar Ismail, K S Tulsi Naik & Rakesh Kumar Mishra

  2. Department of Genetics, University of Delhi, Benito Juarez Marg, South Campus, New Delhi, 110021, India

    Manchikatla Venkat Rajam

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  1. Sahar Ismail
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  2. K S Tulsi Naik
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  3. Manchikatla Venkat Rajam
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Correspondence to K S Tulsi Naik.

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Ismail, S., Tulsi Naik, K.S., Rajam, M.V. et al. Targeting genes involved in nucleopolyhedrovirus DNA multiplication through RNA interference technology to induce resistance against the virus in silkworms. Mol Biol Rep 47, 5333–5342 (2020). https://doi.org/10.1007/s11033-020-05615-z

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