Skip to main content
SpringerLink
Log in

Investigation of Petal Senescence by TRV-Mediated Virus-Induced Gene Silencing in Rose

  • Protocol
  • First Online:
Plant Senescence

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1744))

Abstract

The classic reverse genetic screening, such as EMS-induced or T-DNA-mediated mutation, is a powerful tool to identify senescence-related genes in many model plants. For most non-model plants, however, this strategy is hard to achieve. Even for model plants, construction of a mutant library is usually labor and time-consuming. Virus-induced gene silencing (VIGS) provides an alternative to characterize gene function in a wide spectrum of plants through transient gene expression. To date, more than a dozen of VIGS vector systems have been developed from different RNA and DNA viruses, while Tobacco rattle virus (TRV) system might be one of the most used due to its wide host range and ease of use. Here, we describe a modified TRV vector, TRV-GFP, in which a green fluorescent protein (GFP) is fused to 3′-end of the coat protein (CP) gene in the TRV2 vector. Since the GFP-tagged CP protein could be traced under UV light in planta, identification of TRV-GFP-infected plants is easy. Application of this system in identifying genes regulating petal senescence in rose is described.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Log in via an institution
Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 79.99
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 99.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 129.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

References

  1. Ratcliff F, Martin-Hernandez AM, Baulcombe DC (2001) Technical advance: tobacco rattle virus as a vector for analysis of gene function by silencing. Plant J 25:237–245

    Article  CAS  PubMed  Google Scholar 

  2. Shao Y, Zhu H, Tian H et al (2008) Virus-induced gene silencing in plant species. Russ J Plant Physiol 55:168–174

    Article  CAS  Google Scholar 

  3. Baulcombe D (1999) Viruses and gene silencing in plants. In: 100 years of virology. Springer, New York, pp 189–201

    Chapter  Google Scholar 

  4. Pflieger S, Richard MM, Blanchet S et al (2013) VIGS technology: an attractive tool for functional genomics studies in legumes. Funct Plant Biol 40:1234–1248

    Article  CAS  Google Scholar 

  5. Tian J, Pei H, Zhang S et al (2014) TRV-GFP: a modified Tobacco rattle virus vector for efficient and visualizable analysis of gene function. J Exp Bot 65:311–322

    Article  CAS  PubMed  Google Scholar 

  6. Vaucheret H, Béclin C, Fagard M (2001) Post-transcriptional gene silencing in plants. J Cell Sci 114:3083–3091

    CAS  PubMed  Google Scholar 

  7. Senthil-Kumar M, Mysore KS (2014) Tobacco rattle virus-based virus-induced gene silencing in Nicotiana benthamiana. Nat Protoc 9:1549–1562

    Article  CAS  PubMed  Google Scholar 

  8. Ding S-W, Voinnet O (2007) Antiviral immunity directed by small RNAs. Cell 130:413–426

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Baulcombe D (2004) RNA silencing in plants. Nature 431:356–363

    Article  CAS  PubMed  Google Scholar 

  10. Becker A, Lange M (2010) VIGS-genomics goes functional. Trends Plant Sci 15:1–4

    Article  CAS  PubMed  Google Scholar 

  11. Igarashi A, Yamagata K, Sugai T et al (2009) Apple latent spherical virus vectors for reliable and effective virus-induced gene silencing among a broad range of plants including tobacco, tomato, Arabidopsis thaliana, cucurbits, and legumes. Virology 386:407–416

    Article  CAS  PubMed  Google Scholar 

  12. Scofield SR, Huang L, Brandt AS et al (2005) Development of a virus-induced gene-silencing system for hexaploid wheat and its use in functional analysis of the Lr21-mediated leaf rust resistance pathway. Plant Physiol 138:2165–2173

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Faivre-Rampant O, Gilroy EM, Hrubikova K et al (2004) Potato virus X-induced gene silencing in leaves and tubers of potato. Plant Physiol 134:1308–1316

    Article  CAS  PubMed  PubMed Central  Google Scholar 

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

    Article  CAS  PubMed  Google Scholar 

  15. Liu Y, Schiff M, Marathe R 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  CAS  PubMed  Google Scholar 

  16. Wang C, Cai X, Wang X et al (2006) Optimisation of tobacco rattle virus-induced gene silencing in Arabidopsis. Funct Plant Biol 33:347–355

    Article  CAS  Google Scholar 

  17. Chung E, Seong E, Kim YC et al (2004) A method of high frequency virus induced gene silencing in chili pepper (Capsicum annuum L. cv. Bukang). Mol Cells 17:377–380

    CAS  PubMed  Google Scholar 

  18. Chen J-C, Jiang C-Z, Gookin T et al (2004) Chalcone synthase as a reporter in virus-induced gene silencing studies of flower senescence. Plant Mol Biol 55:521–530

    Article  CAS  PubMed  Google Scholar 

  19. Ma N, Xue J, Li Y et al (2008) Rh-PIP2; 1, a rose aquaporin gene, is involved in ethylene-regulated petal expansion. Plant Physiol 148:894–907

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. Gould B, Kramer EM (2007) Virus-induced gene silencing as a tool for functional analyses in the emerging model plant Aquilegia (columbine, Ranunculaceae). Plant Methods 3:6

    Article  PubMed  PubMed Central  Google Scholar 

  21. Di Stilio VS, Kumar RA, Oddone AM et al (2010) Virus-induced gene silencing as a tool for comparative functional studies in Thalictrum. PLoS One 5:e12064

    Article  PubMed  PubMed Central  Google Scholar 

  22. Jia H, Chai Y, Li C et al (2011) Abscisic acid plays an important role in the regulation of strawberry fruit ripening. Plant Physiol 157:188–199

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Wege S, Scholz A, Gleissberg S et al (2007) Highly efficient virus-induced gene silencing (VIGS) in California poppy (Eschscholzia californica): an evaluation of VIGS as a strategy to obtain functional data from non-model plants. Ann Bot 100:641–649

    Article  PubMed  PubMed Central  Google Scholar 

  24. Ye J, Qu J, Bui HTN et al (2009) Rapid analysis of Jatropha curcas gene functions by virus-induced gene silencing. Plant Biotechnol J 7:964–976

    Article  CAS  PubMed  Google Scholar 

  25. Singh A, Liang Y-C, Kumar P et al (2012) Co-silencing of the Mirabilis antiviral protein (MAP) permits virus-induced gene silencing (VIGS) of other genes in four O’Clock plants (Mirabilis jalapa). J Hortic Sci Biotech 87:334–340

    Article  CAS  Google Scholar 

  26. Gao X, Britt RC Jr, Shan L et al (2011) Agrobacterium-mediated virus-induced gene silencing assay in cotton. J Vis Exp 54:2938

    Google Scholar 

  27. He Z, Chen C (2016) First report of tobacco rattle virus infecting chinese herbaceous peony (Paeonia lactiflora Pall.) in China. Plant Dis 100:2543–2543

    Article  Google Scholar 

  28. Valentine T, Shaw J, Blok VC et al (2004) Efficient virus-induced gene silencing in roots using a modified tobacco rattle virus vector. Plant Physiol 136:3999–4009

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  29. Macfarlane SA (1999) Molecular biology of the tobraviruses. J Gen Virol 80:2799–2807

    Article  CAS  PubMed  Google Scholar 

  30. Lange M, Yellina AL, Orashakova S et al. (2013) Virus-induced gene silencing (VIGS) in plants: an overview of target species and the virus-derived vector systems. Virus-Induced Gene Silencing: Methods and Protocols, 1–14

    Google Scholar 

  31. Liu E, Page JE (2008) Optimized cDNA libraries for virus-induced gene silencing (VIGS) using tobacco rattle virus. Plant Methods 4:5

    Article  PubMed  PubMed Central  Google Scholar 

  32. Xue J, Li Y, Tan H et al (2008) Expression of ethylene biosynthetic and receptor genes in rose floral tissues during ethylene-enhanced flower opening. J Exp Bot 59:2161–2169

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Lü P, Zhang C, Liu J et al (2014) RhHB1 mediates the antagonism of gibberellins to ABA and ethylene during rose (Rosa Hybrida) petal senescence. Plant J 78:578–590

    Article  PubMed  Google Scholar 

  34. Pei H, Ma N, Tian J et al (2013) An NAC transcription factor controls ethylene-regulated cell expansion in flower petals. Plant Physiol 163:775–791

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  35. Wu L, Ma N, Jia Y et al (2016) An ethylene-induced regulatory module delays flower senescence by regulating cytokinin content. Plant Physiol 173:11–21

    Google Scholar 

  36. Senthil-Kumar M, Mysore KS (2011) Virus-induced gene silencing can persist for more than 2 years and also be transmitted to progeny seedlings in Nicotiana benthamiana and tomato. Plant Biotechnol J 9:797–806

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgments

We would thank Dr. Yule Liu (Tsinghua University) for providing the original pTRV vector and for his excellent advices. We would thank Dr. Daqi Fu (China Agricultural University) for his kind and valuable help for the improvement of VIGS approach.

Author information

Authors and Affiliations

  1. Department of Ornamental Horticulture, China Agricultural University, Beijing, China

    Chenxia Cheng, Junping Gao & Nan Ma

  2. Beijing Key Laboratory of Development and Quality Control of Ornamental Crops, China Agricultural University, Beijing, China

    Chenxia Cheng, Junping Gao & Nan Ma

Authors
  1. Chenxia Cheng
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Junping Gao
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Nan Ma
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Nan Ma .

Editor information

Editors and Affiliations

  1. Tobacco Research Institute, Chinese Academy of Agricultural Sciences, Qingdao, Shandong, China

    Yongfeng Guo

Rights and permissions

Reprints and permissions

Copyright information

© 2018 Springer Science+Business Media, LLC

About this protocol

Check for updates. Verify currency and authenticity via CrossMark

Cite this protocol

Cheng, C., Gao, J., Ma, N. (2018). Investigation of Petal Senescence by TRV-Mediated Virus-Induced Gene Silencing in Rose. In: Guo, Y. (eds) Plant Senescence. Methods in Molecular Biology, vol 1744. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-7672-0_4

Download citation

  • DOI: https://doi.org/10.1007/978-1-4939-7672-0_4

  • Published:

  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-7670-6

  • Online ISBN: 978-1-4939-7672-0

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics

Search

Navigation