Abstract
Virus-induced gene silencing (VIGS) is a rapid method for transient silencing of plant genes. In this chapter, we describe the methodology for Tobacco rattle virus (TRV)-based VIGS in Nicotiana tabacum. In combination with subsequent co-expression of the tomato immune receptor Ve1 and the corresponding Verticillium effector Ave1 through Agrobacterium tumefaciens-mediated transient transformation (agroinfiltration), we established a rapid system for assessing the requirement of candidate plant genes for Ve1-mediated immune signaling.
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References
Burch-Smith TM, Anderson JC, Martin GB, Dinesh-Kumar SP (2004) Applications and advantages of virus-induced gene silencing for gene function studies in plants. Plant J 39: 734–746
Burch-Smith TM, Schiff M, Liu YL, Dinesh-Kumar SP (2006) Efficient virus-induced gene silencing in Arabidopsis. Plant Physiol 142: 21–27
Ho FI, Chen YY, Lin YM, Cheng CP, Wang JF (2009) A tobacco rattle virus-induced gene silencing system for a soil-borne vascular pathogen Ralstonia solanacearum. Bot Stud 50:413–424
Liu YL, Schiff M, Dinesh-Kumar SP (2002) Virus-induced gene silencing in tomato. Plant J 31:777–786
Gabriëls SHEJ, Takken FLW, Vossen JH, de Jong CF, Liu Q, Turk SCHJ et al (2006) cDNA-AFLP combined with functional analysis reveals novel genes involved in the hypersensitive response. Mol Plant Microbe Interact 19:567–576
Gabriëls SHEJ, Vossen JH, Ekengren SK, van Ooijen G, Abd-El-Haliem AM, van den Berg GCM et al (2007) An NB-LRR protein required for HR signalling mediated by both extra- and intracellular resistance proteins. Plant J 50:14–28
Fradin EF, Zhang Z, Juarez Ayala JC, Castroverde CD, Nazar RN, Robb J et al (2009) Genetic dissection of Verticillium wilt resistance mediated by tomato Ve1. Plant Physiol 150:320–332
Zhang Z, Fradin EF, de Jonge R, van Esse HP, Smit P, Liu CM et al (2013) Optimized agroinfiltration and virus-induced gene silencing to study Ve1-mediated Verticillium resistance in tobacco. Mol Plant Microbe Interact 26:182–190
Vossen JH, Abd-El-Haliem AM, Fradin EF, van den Berg GCM, Ekengren SK, Meijer HJG et al (2010) Identification of tomato phosphatidylinositol-specific phospholipase-C (PI-PLC) family members and the role of PLC4 and PLC6 in HR and disease resistance. Plant J 62:224–239
Stulemeijer IJE, Stratmann JW, Joosten MHAJ (2007) Tomato mitogen-activated protein kinases LeMPK1, LeMPK2, and LeMPK3 are activated during the Cf-4/Avr4-induced hypersensitive response and have distinct phosphorylation specificities. Plant Physiol 144:1481–1494
Rossi L, Hohn B, Tinland B (1993) The Vird2 protein of Agrobacterium-Tumefaciens carries nuclear-localization signals Important for transfer of T-DNA to plants. Mol Gen Genet 239:345–353
Kapila J, DeRycke R, VanMontagu M, Angenon G (1997) An Agrobacterium-mediated transient gene expression system for intact leaves. Plant Sci 122:101–108
van der Hoorn RAL, Laurent F, Roth R, de Wit PJGM (2000) Agroinfiltration is a versatile tool that facilitates comparative analyses of Avr9/Cf-9-induced and Avr4/Cf-4-induced necrosis. Mol Plant Microbe Interact 13:439–446
de Jonge R, van Esse HP, Maruthachalam K, Bolton MD, Santhanam P, Saber MK et al (2012) Tomato immune receptor Ve1 recognizes effector of multiple fungal pathogens uncovered by genome and RNA sequencing. Proc Natl Acad Sci USA 109:5110–5115
Earley KW, Haag JR, Pontes O, Opper K, Juehne T, Song KM et al (2006) Gateway-compatible vectors for plant functional genomics and proteomics. Plant J 45: 616–629
Fradin EF, Abd-El-Haliem AM, Masini L, van den Berg GCM, Joosten MHAJ, Thomma BPHJ (2011) Interfamily transfer of tomato Ve1 mediates Verticillium resistance in Arabidopsis. Plant Physiol 156:2255–2265
Hu GS, de Hart AKA, Li YS, Ustach C, Handley V, Navarre R et al (2005) EDS1 in tomato is required for resistance mediated by TIR-class R genes and the receptor-like R gene Ve. Plant J 42:376–391
Acknowledgments
We acknowledge Bert Essenstam for excellent plant care. We are grateful to Yin Song and Yan Wang for their help with making the pictures.
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Zhang, Z., Thomma, B.P.H.J. (2014). Virus-Induced Gene Silencing and Agrobacterium tumefaciens-Mediated Transient Expression in Nicotiana tabacum . In: Birch, P., Jones, J., Bos, J. (eds) Plant-Pathogen Interactions. Methods in Molecular Biology, vol 1127. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-986-4_14
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DOI: https://doi.org/10.1007/978-1-62703-986-4_14
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Publisher Name: Humana Press, Totowa, NJ
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Online ISBN: 978-1-62703-986-4
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