Analysis by virus induced gene silencing of the expression of two proline biosynthetic pathway genes in Nicotiana benthamiana under stress conditions

Drought is a major adversity that limits crop yields. Further exploration of wheat drought tolerance-related genes is critical for the genetic improvement of drought tolerance in this crop. Here, comparative proteomic analysis of two wheat varieties, XN979 and LA379, with contrasting drought tolerance was conducted to screen for drought tolerance-related proteins/genes. Virus-induced gene silencing (VIGS) technology was used to verify the functions of candidate proteins. A total of 335 differentially abundant proteins (DAPs) were exclusively identified in the drought-tolerant variety XN979. Most DAPs were mainly involved in photosynthesis, carbon fixation, glyoxylate and dicarboxylate metabolism, and several other pathways. Two DAPs (W5DYH0 and W5ERN8), dubbed TaDrSR1 and TaDrSR2, respectively, were selected for further functional analysis using VIGS. The relative electrolyte leakage rate and malonaldehyde content increased significantly, while the relative water content and proline content significantly decreased in the TaDrSR1- and TaDrSR2-knock-down plants compared to that in non-knocked-down plants under drought stress conditions. TaDrSR1- and TaDrSR2-knock-down plants exhibited more severe drooping and wilting phenotypes than non-knocked-down plants under drought stress conditions, suggesting that the former were more sensitive to drought stress. These results indicate that TaDrSR1 and TaDrSR2 potentially play vital roles in conferring drought tolerance in common wheat.

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Virus-induced gene silencing reveals signal transduction components required for the Pvr9 -mediated hypersensitive response in Nicotiana benthamiana

Virology ◽

10.1016/j.virol.2016.05.011 ◽

2016 ◽

Vol 495 ◽

pp. 167-172 ◽

Cited By ~ 4

Author(s):

Phu-Tri Tran ◽

Hoseong Choi ◽

Doil Choi ◽

Kook-Hyung Kim

Keyword(s):

Signal Transduction ◽

Gene Silencing ◽

Hypersensitive Response ◽

Nicotiana Benthamiana ◽

Virus Induced Gene Silencing

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Virus-induced gene silencing of 14-3-3 genes abrogates dark repression of nitrate reductase activity in Nicotiana benthamiana

Molecular Genetics and Genomics ◽

10.1007/s00438-007-0234-0 ◽

2007 ◽

Vol 278 (2) ◽

pp. 125-133 ◽

Cited By ~ 2

Author(s):

Tatsuya Hirano ◽

Akiko Ito ◽

Thomas Berberich ◽

Ryohei Terauchi ◽

Hiromasa Saitoh

Keyword(s):

Nitrate Reductase ◽

Gene Silencing ◽

Nicotiana Benthamiana ◽

Nitrate Reductase Activity ◽

Reductase Activity ◽

Virus Induced Gene Silencing

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A cucumber green mottle mosaic virus vector for virus-induced gene silencing in cucurbit plants

10.1101/813741 ◽

2019 ◽

Author(s):

Mei Liu ◽

Zhiling Liang ◽

Miguel A. Aranda ◽

Ni Hong ◽

Liming Liu ◽

...

Keyword(s):

Genetic Transformation ◽

Gene Silencing ◽

Mosaic Virus ◽

Gene Function ◽

Nicotiana Benthamiana ◽

Phytoene Desaturase ◽

Virus Induced Gene Silencing ◽

Economic Significance ◽

Potential Alternative ◽

Low Efficiency

AbstractCucurbits produce fruits or vegetables that have great dietary importance and economic significance worldwide. The published genomes of at least 11 cucurbit species are boosting gene mining and novel breeding strategies, however genetic transformation in cucurbits is impractical as a tool for gene function validation due to low transformation efficiencies. Virus-induced gene silencing (VIGS) is a potential alternative tool. So far, very few ideal VIGS vectors are available for cucurbits. Here, we describe a new VIGS vector derived from cucumber green mottle mosaic virus (CGMMV), a monopartite virus that infects cucurbits naturally. We show that the CGMMV vector is competent to induce efficient silencing of the phytoene desaturase (PDS) gene in the model plant Nicotiana benthamiana and in cucurbits, including watermelon, melon, cucumber and bottle gourd. Infection with the CGMMV vector harboring PDS sequences of 69-300 bp in length in the form of sense-oriented or hairpin cDNAs resulted in photobleaching phenotypes in N. benthamiana and cucurbits by PDS silencing. Additional results reflect that silencing of the PDS gene could persist for over two months and the silencing effect of CGMMV-based vectors could be passaged. These results demonstrate that CGMMV vector could serve as a powerful and easy-to-use tool for characterizing gene function in cucurbits.One sentence summaryA CGMMV-based vector enables gene function studies in cucurbits, an extremely low efficiency species for genetic transformation.

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Virus-Induced Gene Silencing in Transgenic Plants Expressing the Minor Capsid Protein of Beet western yellows virus

Molecular Plant-Microbe Interactions ◽

10.1094/mpmi.2002.15.8.799 ◽

2002 ◽

Vol 15 (8) ◽

pp. 799-807 ◽

Cited By ~ 6

Author(s):

V. Brault ◽

S. Pfeffer ◽

M. Erdinger ◽

J. Mutterer ◽

V. Ziegler-Graff

Keyword(s):

Coat Protein ◽

Gene Silencing ◽

Nicotiana Benthamiana ◽

Transcriptional Gene Silencing ◽

Virus Induced Gene Silencing ◽

Minor Capsid Protein ◽

Beet Western Yellows Virus ◽

Post Transcriptional Gene Silencing ◽

Low Levels ◽

Minor Coat Protein

Transgenic Nicotiana benthamiana expressing the minor coat protein P74 of the phloem-limited Beet western yellows virus (BWYV) exhibited an unusual spatial pattern of post-transcriptional gene silencing (PTGS) when infected with BWYV or related viruses. Following infection, transgenic P74 and its mRNA accumulated to only low levels, 21 to 23 nucleotide RNAs homologous to the transgene appeared, and the transgene DNA underwent methylation. The infecting viral RNA, however, was not subject to significant silencing but multiplied readily and produced P74 in the phloem tissues, although the P74 encoded by the transgene disappeared from the phloem as well as the nonvascular tissues.

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Simultaneous silencing of two target genes using virus-induced gene silencing technology in Nicotiana benthamiana

Zeitschrift für Naturforschung C ◽

10.1515/znc-2018-0071 ◽

2019 ◽

Vol 74 (5-6) ◽

pp. 151-159

Author(s):

Feng Zhu ◽

Yanping Che ◽

Fei Xu ◽

Yangkai Zhou ◽

Kun Qian ◽

...

Keyword(s):

Gene Silencing ◽

Nicotiana Benthamiana ◽

Target Genes ◽

Function Analysis ◽

Tobacco Rattle Virus ◽

Pcr Analysis ◽

Virus Induced Gene Silencing ◽

Biotic Stresses ◽

Overlap Extension

Abstract Virus-induced gene silencing (VIGS) is an effective strategy for rapid gene function analysis. It is well established that the NAC transcription factor and salicylic acid (SA) signal pathway play essential roles in response to biotic stresses. However, simultaneous silencing of two target genes using VIGS in plants has been rarely reported. Therefore, in this report, we performed VIGS to silence simultaneously the SA-binding protein 2 (NbSABP2) and NbNAC1 in Nicotiana benthamiana to investigate the gene silencing efficiency of simultaneous silencing of two genes. We first cloned the full-length NbNAC1 gene, and the characterization of NbNAC1 was also analysed. Overlap extension polymerase chain reaction (PCR) analysis showed that the combination of NbSABP2 and NbNAC1 was successfully amplified. Bacteria liquid PCR confirmed that the combination of NbSABP2 and NbNAC1 was successfully inserted into the tobacco rattle virus vector. The results showed that the leaves from the NbSABP2 and NbNAC1 gene-silenced plants collapsed slightly, with browning at the base of petiole or veina. Quantitative real-time PCR results showed that the expression of NbSABP2 and NbNAC1 were significantly reduced in 12 days post silenced plants after tobacco rattle virus infiltration compared with the control plants. Overall, our results suggest that VIGS can be used to silence simultaneously two target genes.

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