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Molecular Biotechnology

, Volume 59, Issue 2–3, pp 73–83 | Cite as

RNAi-Mediated Simultaneous Resistance Against Three RNA Viruses in Potato

  • Amir Hameed
  • Muhammad Nouman Tahir
  • Shaheen Asad
  • Rakhshanda Bilal
  • Joyce Van Eck
  • Georg Jander
  • Shahid Mansoor
Original Paper

Abstract

RNA interference (RNAi) technology has been successfully applied in stacking resistance against viruses in numerous crop plants. During RNAi, the production of small interfering RNAs (siRNAs) from template double-standard RNA (dsRNA) derived from expression constructs provides an on-switch for triggering homology-based targeting of cognate viral transcripts, hence generating a pre-programmed immunity in transgenic plants prior to virus infection. In the current study, transgenic potato lines (Solanum tuberosum cv. Desiree) were generated, expressing fused viral coat protein coding sequences from Potato virus X (PVX), Potato virus Y (PVY), and Potato virus S (PVS) as a 600-bp inverted repeat expressed from a constitutive 35S promoter. The expression cassette (designated Ec1/p5941) was designed to generate dsRNAs having a hairpin loop configuration. The transgene insertions were confirmed by glufosinate resistance, gene-specific PCR, and Southern blotting. Regenerated lines were further assayed for resistance to virus inoculation for up to two consecutive crop seasons. Nearly 100% resistance against PVX, PVY, and PVS infection was observed in transgenic lines when compared with untransformed controls, which developed severe viral disease symptoms. These results establish the efficacy of RNAi using the coat protein gene as a potential target for the successful induction of stable antiviral immunity in potatoes.

Keywords

RNAi Interaction Resistance Virus Transgenic Potato PVX PVY PVS 

Notes

Acknowledgements

This work was funded by Higher Education Commission (HEC), Government of Pakistan, under the indigenous PhD fellowship and research initiative program (IRSIP), and United States Department of Agriculture award 2014-67013-21659 to GJ. The authors would like to thank Ms. Kerry Swartwood for the assistance in tissue culture work conducted at the Boyce Thomason Institute for Plant Research.

Compliance with Ethical Standards

Conflict of interest

The authors declare no financial or commercial conflict of interest.

Supplementary material

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Supplementary material 1 (DOCX 13 kb)
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Supplementary material 2 (DOCX 13 kb)
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Supplementary material 3 (DOCX 12 kb)
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Supplementary Fig. 1 Resistance analysis of transgenic plants (T0 generation, second-year crop). a Photograph taken 10 days of postemergence of transgenic plants shows healthy appearance with no disease symptoms anywhere. b Untransformed control plant showing viral disease symptoms of leaf thickening and greening at 10 days postemergence. c Transgenic plant showing healthy growth at 30 days postemergence. d Untransformed control plant showing severe viral disease symptoms of leaf curling, shrinking, mosaic and stunted growth at 30 days postemergence. (JPEG 1599 kb)

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Copyright information

© Springer Science+Business Media New York 2017

Authors and Affiliations

  • Amir Hameed
    • 1
    • 2
  • Muhammad Nouman Tahir
    • 1
  • Shaheen Asad
    • 1
  • Rakhshanda Bilal
    • 1
  • Joyce Van Eck
    • 3
  • Georg Jander
    • 3
  • Shahid Mansoor
    • 1
  1. 1.Molecular Virology and Gene Silencing GroupNational Institute for Biotechnology and Genetic EngineeringFaisalabadPakistan
  2. 2.Department of Bioinformatics and BiotechnologyGovernment College UniversityFaisalabadPakistan
  3. 3.Boyce Thompson Institute for Plant ResearchIthacaUSA

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