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Virus-Induced Gene Silencing of N Gene in Tobacco by Apple Latent Spherical Virus Vectors

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Plant Virology Protocols

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1236))

Abstract

Virus infections induce an RNA-mediated defense that targets viral RNAs in a nucleotide sequence-specific manner in plants, commonly referred to as virus-induced gene silencing (VIGS). When the virus carries sequences of plant genes, it triggers virus-induced gene silencing (VIGS) and results in the degradation of mRNA of endogenous homologous gene. VIGS has been shown to have great potential as a reverse-genetics tool for studying of gene functions in plants, and it has several advantages over other functional genomics approaches. Here, we describe VIGS of N gene in tobacco cv. Xanthi nc by ALSV vectors containing fragments of N gene from Nicotiana glutinosa.

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References

  1. Lu R et al (2003) Virus-induced gene silencing in plants. Methods 30:296–303

    Article  PubMed  CAS  Google Scholar 

  2. Robertson D (2004) VIGS vector for gene silencing: many targets, many tools. Annu Rev Plant Biol 55:495–519

    Article  PubMed  CAS  Google Scholar 

  3. Purkayastha A, Dasgupta I (2009) Virus-induced gene silencing: a versatile tool for discovery of gene functions in plants. Plant Physiol Biochem 47:967–976

    Article  PubMed  CAS  Google Scholar 

  4. Goodin MM et al (2008) Nicotiana benthamiana: its history and future as a model for plant-pathogen interactions. Mol Plant-Microbe Interact 21:1015–1026

    Article  PubMed  CAS  Google Scholar 

  5. Burch-Smith TM et al (2004) Applications and advantages of virus-induced gene silencing for gene function studies in plants. Plant J 39: 734–746

    Article  PubMed  CAS  Google Scholar 

  6. Hofgen R et al (1994) A visible marker for antisense mRNA expression in plants: inhibition of chlorophyll synthesis with a glutamate-1-semialdehyde aminotransferase antisense gene. Proc Natl Acad Sci U S A 91:1726–1730

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  7. Romeis T et al (2001) Calcium-dependent protein kinases play an essential role in a plant defence response. EMBO J 20:5556–5567

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  8. Hiriart JB et al (2002) Suppression of a key gene involved in chlorophyll biosynthesis by means of virus-inducing gene silencing. Plant Mol Biol 50:213–224

    Article  PubMed  CAS  Google Scholar 

  9. Liu Y, Schiff M, Dinesh-Kumar SP (2002) Virus-induced gene silencing in tomato. Plant J 31:777–786

    Article  PubMed  CAS  Google Scholar 

  10. Cai X-Z et al (2006) Development of a virus-induced gene-silencing system for functional analysis of the RPS2-dependent resistance signalling pathways in Arabidopsis. Plant Mol Biol 62:223–232

    Article  PubMed  CAS  Google Scholar 

  11. Deng X et al (2012) Virus-induced gene silencing for Asteraceae - a reverse genetics approach for functional genomics in Gerbera hybrid. Plant Biotechnol J 10:970–978

    Article  PubMed  CAS  Google Scholar 

  12. Manmathan H et al (2013) Virus-induced gene silencing of Arabidopsis thaliana gene homologues in wheat identifies genes conferring improved drought tolerance. J Exp Bot 64:1381–1392

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  13. Kang B, Yeam I, Jahn MM (2005) Genetics of plant virus resistance. Annu Rev Phytopathol 43:581–621

    Article  PubMed  CAS  Google Scholar 

  14. Whitham S et al (1994) The product of the tobacco mosaic virus resistance gene N: similarity to Toll and the interleukin-1 receptor. Cell 78:1101–1115

    Article  PubMed  CAS  Google Scholar 

  15. Liu Y et al (2002) Tobacco Rar1, EDS1 and NPR1/NIM1 like genes are required for N-mediated resistance to tobacco mosaic virus. Plant J 30:415–429

    Article  PubMed  CAS  Google Scholar 

  16. Li C et al (2000) Nucleotide sequence and genome organization of Apple latent spherical virus: a new virus classified into the family Comoviridae. J Gen Virol 81:541–547

    PubMed  CAS  Google Scholar 

  17. Li C et al (2004) Stable expression of foreign proteins in herbaceous and apple plants using Apple latent spherical virus RNA2 vectors. Arch Virol 149:1541–1558

    PubMed  CAS  Google Scholar 

  18. Takahashi T, Yoshikawa N (2008) Analysis of cell-to-cell and long-distance movement of Apple latent spherical virus in infected plants using green, cyan, and yellow fluorescent proteins. In: Forester G, Johansen E, Hong Y, Nagy P (eds) Plant virology protocols, vol 451, Methods in molecular biology. Humana Press, Totowa, NJ, pp 545–554

    Chapter  Google Scholar 

  19. Yaegashi H et al (2007) Characterization of virus-induced gene silencing in tobacco plants infected with apple latent spherical virus. Arch Virol 152:1839–1849

    Article  PubMed  CAS  Google Scholar 

  20. Igarashi A et al (2009) Apple latent spherical virus vectors for reliable and effective virusinduced gene silencing among a broad range of plants infecting tobacco, tomato, Arabidopsis thaliana, cucurbits, and legumes. Virology 386:407–416

    Article  PubMed  CAS  Google Scholar 

  21. Yamagishi N, Yoshikawa N (2009) Virus-induced gene silencing in soybean seeds and the emergence of soybean plants with Apple latent spherical virus vectors. Plant Mol Biol 71:15–24

    Article  PubMed  CAS  Google Scholar 

  22. Yamagishi N et al (2011) Promotion of flowering and reduction of a generation time in apple seedlings by ectopical expression of the Arabidopsis thaliana FT gene using the Apple latent spherical virus vector. Plant Mol Biol 75:193–204

    Article  PubMed  CAS  Google Scholar 

  23. Yamagishi N, Kishigami R, Yoshikawa N (2014) Reduced generation time of apple seedlings to within a year by means of a plant virus vector: a new plant-breeding technique with no transmission of genetic modification to the next generation. Plant Biotechnol J 12:60

    Article  PubMed  CAS  Google Scholar 

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Correspondence to Nobuyuki Yoshikawa .

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Li, C., Yoshikawa, N. (2015). Virus-Induced Gene Silencing of N Gene in Tobacco by Apple Latent Spherical Virus Vectors. In: Uyeda, I., Masuta, C. (eds) Plant Virology Protocols. Methods in Molecular Biology, vol 1236. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-1743-3_17

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  • DOI: https://doi.org/10.1007/978-1-4939-1743-3_17

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-1742-6

  • Online ISBN: 978-1-4939-1743-3

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