TRV–GFP: a modified Tobacco rattle virus vector for efficient and visualizable analysis of gene function

Tian, Ji; Pei, Haixia; Zhang, Shuai; Chen, Jiwei; Chen, Wen; Yang, Ruoyun; Meng, Yonglu; You, Jun; Gao, Junping; Ma, Nan

doi:10.1093/jxb/ert381

Cited by 139 publications

(110 citation statements)

References 38 publications

(52 reference statements)

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“…Silencing of RhPR10.1 and RhHB6 in petal discs and plantlets by VIGS was conducted as described previously Dai et al, 2012;Tian et al, 2014), but with some modifications. The pTRV1, pTRV2, pTRV-RhPR10.1, and pTRV-RhHB6 vectors were transformed into Agrobacterium tumefaciens strain GV3101.…”

Section: Vigsmentioning

confidence: 99%

An Ethylene-Induced Regulatory Module Delays Flower Senescence by Regulating Cytokinin Content

Jia

et al. 2016

Plant Physiol.

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103

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ORCID IDs: 0000-0002-6288-7917 (Y.J.); 0000-0002-5972-7963 (C.-Z.J.); 0000-0002-3866-0894 (C.M.); 0000-0002-9285-2539 (J.G.).In many plant species, including rose (Rosa hybrida), flower senescence is promoted by the gaseous hormone ethylene and inhibited by the cytokinin (CTK) class of hormones. However, the molecular mechanisms underlying these antagonistic effects are not well understood. In this study, we characterized the association between a pathogenesis-related PR-10 family gene from rose (RhPR10.1) and the hormonal regulation of flower senescence. Quantitative reverse transcription PCR analysis showed that RhPR10.1 was expressed at high levels during senescence in different floral organs, including petal, sepal, receptacle, stamen, and pistil, and that expression was induced by ethylene treatment. Silencing of RhPR10.1 expression in rose plants by virusinduced gene silencing accelerated flower senescence, which was accompanied by a higher ion leakage rate in the petals, as well as increased expression of the senescence marker gene RhSAG12. CTK content and the expression of three CTK signaling pathway genes were reduced in RhPR10.1-silenced plants, and the accelerated rate of petal senescence that was apparent in the RhPR10.1-silenced plants was restored to normal levels by CTK treatment. Finally, RhHB6, a homeodomain-Leu zipper I transcription factor, was observed to bind to the RhPR10.1 promoter, and silencing of its expression also promoted flower senescence. Our results reveal an ethylene-induced RhHB6-RhPR10.1 regulatory module that functions as a brake of ethylenepromoted senescence through increasing the CTK content.

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Section: Vigsmentioning

confidence: 99%

An Ethylene-Induced Regulatory Module Delays Flower Senescence by Regulating Cytokinin Content

Jia

et al. 2016

Plant Physiol.

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103

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“…In VIGS systems, silencing phenotypes often appear 3-4 weeks after infection, but can sometimes appear at >6 weeks after inoculation (Di Stilio et al 2010;Tian et al 2014;Zhong et al 2014). In this study, N. benthamiana plants exhibited the photo-bleaching phenotype at approximately 17 dai (Figure 2).…”

Section: Discussionmentioning

confidence: 54%

<i>Peanut stunt virus</i>-induced gene silencing in white lupin (<i>Lupinus albus</i>)

Yamagishi

Masuta

Suzuki

et al. 2015

Plant Biotechnology

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White lupin (Lupinus albus L.) plants develop cluster roots and have strong resistance to phosphorus starvation. Although many expressed sequences have been identified to explain the mechanisms used by white lupin to acquire phosphorus, the lack of a stable transformation technique has made it challenging to evaluate the functions of these expressed sequences. Virus-induced gene silencing (VIGS) is an attractive method for assaying gene function in species that are difficult to stably transform. Here, we show that the Peanut stunt virus (PSV) vector effectively induces silencing of endogenous genes in white lupin. It is unknown whether PSV is useful for VIGS; therefore, we first inoculated Nicotiana benthamiana plants with PSV harbouring fragments of the N. benthamiana phytoene desaturase gene (NbPDS). Two out of four distinct sequence fragments of NbPDS induced photo-bleaching in N. benthamiana, indicating that PSV can be used to knockdown endogenous gene sequences in a sequence-dependent manner. White lupin plants inoculated with PSV harbouring fragments of the L. albus PDS gene (LaPDS) developed photo-bleaching that was associated with a significant reduction in LaPDS mRNA accumulation. PSV spread systemically in leaves, roots, and cluster roots, and small interfering RNA of LaPDS was detected in these organs. This is the first study to demonstrate the use of VIGS by PSV, suggesting that this vector can be applied to supress endogenous gene expression in shoots and roots of white lupin and to clarify the mechanisms of phosphorus starvation resistance.

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“…A modified Tobacco rattle virus-GFP vector has been also produced recently for efficient and visualizable analysis of gene function. 19 For a GLRaV-2-derived vector, systemic gene expression was detectable in grapevine from 4weeks post inoculation.…”

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confidence: 99%

Agrobacterium and Viral Vectors-Mediated in vivo Gene Silencing Assays in Plants

Ben-Amar¹

2017

OAJS

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Over the last years, the explosive growth of plant genomic resources and the development of advanced vectors enhanced the emergence of rapid systems to dissect plant gene function. Although transgenic approach has been applied to generate stable transgenic lines, this procedure is time-consuming and labor-intensive. Transient expression systems are continuously emerging during these years as an alternative tool for functional genomics studies in plants. Among these techniques, Agrobacterium-infiltration and infectious clones-derived plant viruses have been developed and successfully applied for versatile transient assays. Their effectiveness over a wide range of plant species is often validated. Here, the green fluorescent protein (GFP) gene was investigated as reporter gene for proof-of-concept to induce gene silencing in GFP-transgenic tobacco and grapevine plants either by inoculation of Arabis mosaic virus infectious clones or through in planta vacuum infiltration of Agrobacterium suspension.Leaves derived from greenhouse-grown grapevine plants were vacuum infiltrated with agrobacteria suspension containing GFP gene to induce silencing. Nicotiana benthamiana plants were agro-inoculated with Arabis mosaic virus infectious clones RNA 1 and RNA 2 along with a modified RNA satellite harboring a GFP construct. Systemic GFP-silencing was monitored during days and weeks after inoculation and data showed that silencing was achieved after 10-12 days post inoculation for tobacco, while it takes 6 weeks to be completely reached in grapevine leaves. The transient expression systems established here were successfully applied for inducing a knock-out of endogenous gene. These techniques provide a fast and reliable tool to carry out gene function analysis in plants and open a future outlook with several downstream applications towards pathogen resistance or abiotic stress tolerance.

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TRV–GFP: a modified Tobacco rattle virus vector for efficient and visualizable analysis of gene function

Abstract: We developed an easy-traceable TRV vector, TRV2-GFP, by tagging a GFP to the coat protein. TRV2-GFP-infected plants could be identified efficiently by GFP monitoring. TRV2-GFP is useful for functional genomics in many plants, especially for non-Solanaceae plants, like rose

Cited by 139 publications

References 38 publications

An Ethylene-Induced Regulatory Module Delays Flower Senescence by Regulating Cytokinin Content

An Ethylene-Induced Regulatory Module Delays Flower Senescence by Regulating Cytokinin Content

<i>Peanut stunt virus</i>-induced gene silencing in white lupin (<i>Lupinus albus</i>)

Agrobacterium and Viral Vectors-Mediated in vivo Gene Silencing Assays in Plants

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