High rates of virus-induced gene silencing by tobacco rattle virus in Populus
2015 | journal article. A publication with affiliation to the University of Göttingen.
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High rates of virus-induced gene silencing by tobacco rattle virus in Populus
Shen, Z.; Sun, J.; Yao, J.; Wang, S.; Ding, M.; Zhang, H. & Qian, Z. et al. (2015)
Tree Physiology, 35(9) pp. 1016-1029. DOI: https://doi.org/10.1093/treephys/tpv064
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- Authors
- Shen, Zedan; Sun, Jian; Yao, Jun; Wang, Shaojie; Ding, Mingquan; Zhang, Huilong; Qian, Zeyong; Zhao, Nan; Sa, Gang; Zhao, Rui; Shen, Xin; Polle, Andrea ; Chen, Shaoliang
- Abstract
- Virus-induced gene silencing (VIGS) has been shown to be an effective tool for investigating gene functions in herbaceous plant species, but has rarely been tested in trees. The establishment of a fast and reliable transformation system is especially important for woody plants, many of which are recalcitrant to transformation. In this study, we established a tobacco rattle virus (TRV)-based VIGS system for two Populus species, Populus euphratica and P. × canescens. Here, TRV constructs carrying a 266 bp or a 558 bp fragment of the phytoene desaturase (PDS) gene were Agrobacterium-infiltrated into leaves of the two poplar species. Agrobacterium-mediated delivery of the shorter insert, TRV2-PePDS266, into the host poplars resulted in expected photobleaching in both tree species, but not the longer insert, PePDS558. The efficiency of VIGS was temperature-dependent, increasing by raising the temperature from 18 to 28 °C. The optimized TRV–VIGS system at 28 °C resulted in a high silencing frequency and efficiency up to 65–73 and 83–94%, respectively, in the two tested poplars. Moreover, syringe inoculation of Agrobacterium in 100 mM acetosyringone induced a more efficient silencing in the two poplar species, compared with other agroinfiltration methods, e.g., direct injection, misting and agrodrench. There were plant species-related differences in the response to VIGS because the photobleaching symptoms were more severe in P. × canescens than in P. euphratica. Furthermore, VIGS-treated P. euphratica exhibited a higher recovery rate (50%) after several weeks of the virus infection, compared with TRV-infected P. × canescens plants (20%). Expression stability of reference genes was screened to assess the relative abundance of PePDS mRNA in VIGS-treated P. euphratica and P. × canescens. PeACT7 was stably expressed in P. euphratica and UBQ-L was selected as the most suitable reference gene for P. × canescens using three different statistical approaches, geNorm, NormFinder and BestKeeper. Quantitative real-time PCR showed significant reductions in PDS transcripts (55–64%) in the photobleached leaves of both VIGS-treated poplar species. Our results demonstrate that the TRV-based VIGS provides a practical tool for gene functional analysis in Populus sp., especially in those poplar species which are otherwise recalcitrant to transformation.
- Issue Date
- 2015
- Journal
- Tree Physiology
- Organization
- Fakultät für Forstwissenschaften und Waldökologie ; Büsgen-Institut ; Abteilung Forstbotanik und Baumphysiologie
- ISSN
- 0829-318X
- Language
- English