Establishment of a Virus-Induced Gene-Silencing (VIGS) System in Tea Plant and Its Use in the Functional Analysis of <i>CsTCS1</i>
Tea (<i>Camellia sinensis</i> [L.] O. Kuntze) is an important global economic crop and is considered to enhance health. However, the functions of many genes in tea plants are unknown. Virus-induced gene silencing (VIGS) mediated by tobacco rattle virus (TRV) is an effective tool for the...
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| Format: | Article |
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MDPI AG
2022-12-01
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| Series: | International Journal of Molecular Sciences |
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| Online Access: | https://www.mdpi.com/1422-0067/24/1/392 |
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| author | Guodong Li Yan Li Xinzhuan Yao Litang Lu |
| author_facet | Guodong Li Yan Li Xinzhuan Yao Litang Lu |
| author_sort | Guodong Li |
| collection | DOAJ |
| description | Tea (<i>Camellia sinensis</i> [L.] O. Kuntze) is an important global economic crop and is considered to enhance health. However, the functions of many genes in tea plants are unknown. Virus-induced gene silencing (VIGS) mediated by tobacco rattle virus (TRV) is an effective tool for the analysis of gene functions, although this method has rarely been reported in tea plants. In this study, we established an effective VIGS-mediated gene knockout technology to understand the functional identification of large-scale genomic sequences in tea plants. The results showed that the VIGS system was verified by detecting the virus and using a real-time quantitative reverse transcription PCR (qRT-PCR) analysis. The reporter gene <i>CsPOR1</i> (protochlorophyllide oxidoreductase) was silenced using the vacuum infiltration method, and typical photobleaching and albino symptoms were observed in newly sprouted leaves at the whole plant level of tea after infection for 12 d and 25 d. After optimization, the VIGS system was successfully used to silence the tea plant <i>CsTCS1</i> (caffeine synthase) gene. The results showed that the relative caffeine content was reduced 6.26-fold compared with the control, and the level of expression of <i>CsPOR1</i> decreased by approximately 3.12-fold in plants in which <i>CsPOR1</i> was silenced. These results demonstrate that VIGS can be quickly and efficiently used to analyze the function of genes in tea plants. The successful establishment of VIGS could eliminate the need for tissue culture by providing an effective method to study gene function in tea plants and accelerate the process of functional genome research in tea. |
| first_indexed | 2024-03-11T09:59:50Z |
| format | Article |
| id | doaj.art-6bf65a429d0c4de9810887b864b53a25 |
| institution | Directory Open Access Journal |
| issn | 1661-6596 1422-0067 |
| language | English |
| last_indexed | 2024-03-11T09:59:50Z |
| publishDate | 2022-12-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | International Journal of Molecular Sciences |
| spelling | doaj.art-6bf65a429d0c4de9810887b864b53a252023-11-16T15:32:44ZengMDPI AGInternational Journal of Molecular Sciences1661-65961422-00672022-12-0124139210.3390/ijms24010392Establishment of a Virus-Induced Gene-Silencing (VIGS) System in Tea Plant and Its Use in the Functional Analysis of <i>CsTCS1</i>Guodong Li0Yan Li1Xinzhuan Yao2Litang Lu3The Key Laboratory of Plant Resources Conservation and Germplasm Innovation in Mountainous Region (Ministry of Education), College of Life Sciences, Guizhou University, Guiyang 550025, ChinaThe Key Laboratory of Plant Resources Conservation and Germplasm Innovation in Mountainous Region (Ministry of Education), College of Life Sciences, Guizhou University, Guiyang 550025, ChinaThe Key Laboratory of Plant Resources Conservation and Germplasm Innovation in Mountainous Region (Ministry of Education), College of Life Sciences, Guizhou University, Guiyang 550025, ChinaThe Key Laboratory of Plant Resources Conservation and Germplasm Innovation in Mountainous Region (Ministry of Education), College of Life Sciences, Guizhou University, Guiyang 550025, ChinaTea (<i>Camellia sinensis</i> [L.] O. Kuntze) is an important global economic crop and is considered to enhance health. However, the functions of many genes in tea plants are unknown. Virus-induced gene silencing (VIGS) mediated by tobacco rattle virus (TRV) is an effective tool for the analysis of gene functions, although this method has rarely been reported in tea plants. In this study, we established an effective VIGS-mediated gene knockout technology to understand the functional identification of large-scale genomic sequences in tea plants. The results showed that the VIGS system was verified by detecting the virus and using a real-time quantitative reverse transcription PCR (qRT-PCR) analysis. The reporter gene <i>CsPOR1</i> (protochlorophyllide oxidoreductase) was silenced using the vacuum infiltration method, and typical photobleaching and albino symptoms were observed in newly sprouted leaves at the whole plant level of tea after infection for 12 d and 25 d. After optimization, the VIGS system was successfully used to silence the tea plant <i>CsTCS1</i> (caffeine synthase) gene. The results showed that the relative caffeine content was reduced 6.26-fold compared with the control, and the level of expression of <i>CsPOR1</i> decreased by approximately 3.12-fold in plants in which <i>CsPOR1</i> was silenced. These results demonstrate that VIGS can be quickly and efficiently used to analyze the function of genes in tea plants. The successful establishment of VIGS could eliminate the need for tissue culture by providing an effective method to study gene function in tea plants and accelerate the process of functional genome research in tea.https://www.mdpi.com/1422-0067/24/1/392tea plantvirus-induced gene silencing (VIGS)tobacco rattle virus (TRV)<i>CsPOR1</i><i>CsTCS1</i> |
| spellingShingle | Guodong Li Yan Li Xinzhuan Yao Litang Lu Establishment of a Virus-Induced Gene-Silencing (VIGS) System in Tea Plant and Its Use in the Functional Analysis of <i>CsTCS1</i> International Journal of Molecular Sciences tea plant virus-induced gene silencing (VIGS) tobacco rattle virus (TRV) <i>CsPOR1</i> <i>CsTCS1</i> |
| title | Establishment of a Virus-Induced Gene-Silencing (VIGS) System in Tea Plant and Its Use in the Functional Analysis of <i>CsTCS1</i> |
| title_full | Establishment of a Virus-Induced Gene-Silencing (VIGS) System in Tea Plant and Its Use in the Functional Analysis of <i>CsTCS1</i> |
| title_fullStr | Establishment of a Virus-Induced Gene-Silencing (VIGS) System in Tea Plant and Its Use in the Functional Analysis of <i>CsTCS1</i> |
| title_full_unstemmed | Establishment of a Virus-Induced Gene-Silencing (VIGS) System in Tea Plant and Its Use in the Functional Analysis of <i>CsTCS1</i> |
| title_short | Establishment of a Virus-Induced Gene-Silencing (VIGS) System in Tea Plant and Its Use in the Functional Analysis of <i>CsTCS1</i> |
| title_sort | establishment of a virus induced gene silencing vigs system in tea plant and its use in the functional analysis of i cstcs1 i |
| topic | tea plant virus-induced gene silencing (VIGS) tobacco rattle virus (TRV) <i>CsPOR1</i> <i>CsTCS1</i> |
| url | https://www.mdpi.com/1422-0067/24/1/392 |
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