Abstract
Virus infection leads to the synthesis of double-stranded RNA during virus replication, and then this infection produces small RNA molecules in the antiviral RNA silencing pathway. Here, we develop an Agrobacterium-mediated inoculation system for ALSV-based vectors. This system is more effective and convenient for inoculation of the ALSV vectors into plants compared to direct inoculation of ALSV-RNA2-based vectors in pUC plasmids reported previously. In addition, cointroduction of various plant viral RNA-silencing suppressors increased the efficiency of agroinoculation of the ALSV-based vector. An ALSV vector could be successfully used to silence an endogenous gene in plants. Therefore, the ALSV-based VIGS/agroinoculation system provides a valuable tool for functional genomics among a broad range of plant species.
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References
Kon T, Ikegami M (2009) RNA silencing in plants and its suppression by plant viruses. CAB Reviews 4(64):1–16
Aliyari R, Ding SW (2009) RNA-based viral immunity initiated by the Dicer family of host immune receptors. Immunol Rev 227:176–188
Vazquez F (2006) Arabidopsis endogenous small RNAs: Highways and byways. Trends Plant Sci 11:460–468
Senthil-Kumar M, Mysore KS (2011) New dimensions for VIGS in plant functional genomics. Trends Plant Sci 16:656–665
Le Gall O, Sanfaçon H, Ikegami M et al (2007) Cheravirus and Sadwavirus: two unassigned genera of plant positive-sense single-stranded RNA viruses formerly considered atypical members of the genus Nepovirus (family Comoviridae). Arch Virol 152:1767–1774
Li C, Yoshikawa N, Takahashi T 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
Li C, Sasaki N, Isogai M 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
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
Vaghchhipawala Z, Rojas CM, Senthil-Kumar M et al (2011) Agroinoculation and agroinfiltration: simple tools for complex gene function analyses. Methods Mol Biol 678:65–76
Hajdukiewicz P, Svab Z, Maliga P (1994) The small, versatile pPZP family of Agrobacterium binary vectors for plant transformation. Plant Mol Biol 25:989–994
Singh K, Raizada J, Bhardwaj P et al (2004) 26S rRNA-based internal control gene primer pair for reverse transcription-polymerase chain reaction-based quantitative expression studies in diverse plant species. Anal Biochem 335:330–333
Acknowledgments
This work was supported by the Program for Promotion of Basic and Applied Research for Innovations in Bio-oriented Industry.
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Kon, T., Yoshikawa, N. (2015). An Effective and Convenient Method for the Delivery of Apple Latent Spherical Virus (ALSV)-Based Vectors into Plant Cells by Agroinoculation. In: Mysore, K., Senthil-Kumar, M. (eds) Plant Gene Silencing. Methods in Molecular Biology, vol 1287. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-2453-0_14
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DOI: https://doi.org/10.1007/978-1-4939-2453-0_14
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