Development of Agrobacterium-Mediated Virus-Induced Gene Silencing and Performance Evaluation of Four Marker Genes in Gossypium barbadense

Pang, Junfeng; Zhu, Yue; Li, Qing; Jin-zhi, Liu; Tian, Yingchuan; Liu, Yule; Wu, Jiahe

doi:10.1371/journal.pone.0073211

Cited by 71 publications

(63 citation statements)

References 36 publications

(63 reference statements)

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“…In a preliminary experiment, we silenced the expression of the phytoene desaturase gene (GbPDS), which is used as a visible marker to monitor the efficiency of VIGS (Pang et al, 2013). As shown in Figure 7B, the newly emerged true leaves of plants infiltrated with Agrobacterium carrying GbPDS exhibited an albino phenotype compared to the negative control (Fig.…”

Section: Silencing Of Nrx1 Leads To Reduced V Dahliae Resistancementioning

confidence: 99%

The Thioredoxin GbNRX1 Plays a Crucial Role in Homeostasis of Apoplastic Reactive Oxygen Species in Response to Verticillium dahliae Infection in Cotton

et al. 2016

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Examining the proteins that plants secrete into the apoplast in response to pathogen attack provides crucial information for understanding the molecular mechanisms underlying plant innate immunity. In this study, we analyzed the changes in the root apoplast secretome of the Verticillium wilt-resistant island cotton cv Hai 7124 (Gossypium barbadense) upon infection with Verticillium dahliae. Two-dimensional differential gel electrophoresis and matrix-assisted laser desorption/ionization tandem time-of-flight mass spectrometry analysis identified 68 significantly altered spots, corresponding to 49 different proteins. Gene ontology annotation indicated that most of these proteins function in reactive oxygen species (ROS) metabolism and defense response. Of the ROS-related proteins identified, we further characterized a thioredoxin, GbNRX1, which increased in abundance in response to V. dahliae challenge, finding that GbNRX1 functions in apoplastic ROS scavenging after the ROS burst that occurs upon recognition of V. dahliae. Silencing of GbNRX1 resulted in defective dissipation of apoplastic ROS, which led to higher ROS accumulation in protoplasts. As a result, the GbNRX1-silenced plants showed reduced wilt resistance, indicating that the initial defense response in the root apoplast requires the antioxidant activity of GbNRX1. Together, our results demonstrate that apoplastic ROS generation and scavenging occur in tandem in response to pathogen attack; also, the rapid balancing of redox to maintain homeostasis after the ROS burst, which involves GbNRX1, is critical for the apoplastic immune response.

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Section: Silencing Of Nrx1 Leads To Reduced V Dahliae Resistancementioning

confidence: 99%

The Thioredoxin GbNRX1 Plays a Crucial Role in Homeostasis of Apoplastic Reactive Oxygen Species in Response to Verticillium dahliae Infection in Cotton

et al. 2016

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“…For the virus-induced gene silencing (VIGS) analysis, tobacco rattle virus ( TRV )-based vectors, including pTRV1 (pYL192), pTRV2 (pYL156) and pTRV2e were used in this study. TRV:PDS was employed as a positive control vector in the silenced plants, which had been previously reported by Pang et al (2013). The construction of the TRV-related vectors was performed as described by Liu et al (2004) and Sha et al (2014).…”

Section: Methodsmentioning

confidence: 99%

The ghr-miR164 and GhNAC100 module participates in cotton plant defence against Verticillium dahliae

Wang

et al. 2018

Preprint

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Previous reports have shown that many miRNAs were identified at the early induction stage during which Verticillium dahliae localizes at the root surface. In this study, we constructed two sRNA libraries of cotton root responses to this fungus at the later induction stage when the pathogen enters the root vascular tissue. We identified 71 known miRNAs and 378 novel miRNAs from two pathogen-induced sRNAs and the control libraries. Combined with degradome and sRNA sequencing, 178 corresponding miRNA target genes were identified, in which 40 target genes from differentially expressed miRNAs were primarily associated with oxidation-reduction and stress responses. More importantly, we characterized the ghr-miR164-GhNAC100 module in the response of the plant to V dahliae infection. A GUS fusion reporter showed that ghr-miR164 directly cleaved the mRNA of GhNAC100 in the post-transcriptional process. ghr-miR164-silencing increased the resistance of the plant to this fungus, while the knockdown of GhNAC100 elevated the susceptibility of the plant, indicating that ghr-miR164-GhNAC100 modulates plant defence through the post-transcriptional regulation. Our data documented that there are numerous miRNAs at the later induction stage that participate in the plant response to V. dahliae, suggesting that miRNAs play important roles in plant resistance to vascular disease.HighlightAccording to degradome and sRNA sequencings of cotton root in responses to Verticillium dahliae at the later induction stage, many miRNAs and corresponding targets including ghr-miR164-GhNAC100 module participate plant defence.

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“…Tobacco rattle virus, TRV1 and TRV2 (pYL156), were used as vectors for the VIGS (Virus-induced gene silencing) assay. The cotton phytoene desaturase (PDS) was used to check the efficiency of VIGS (Tuttle et al 2008;Pang et al 2013), since the silencing of PDS gene caused the loss of carotenoids and chlorophyll which resulting in white leaves. The pYL156 was employed as the negative control.…”

Section: Vigs (Virus-induced Gene Silencing) Assaymentioning

confidence: 99%

Map-based cloning of a recessive gene v1 for virescent leaf expression in cotton (Gossypium spp.)

et al. 2018

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Background: Virescence, as a recognizable phenotype in the early development stage of cotton, is not only available for research on chloroplast development and photosynthesis but also for heterosis exploitation in cotton. Methods: In current study, for fine mapping of virescent-1 (v 1 ) in cotton, three populations with a total of 5 678 individuals were constructed using T582 which has the virescent trait. Tobacco rattle virus, TRV1 and TRV2 (pYL156), were used as vectors for the virus-induced gene silencing (VIGS) assay. Results:The v 1 gene was fine-mapped to a 20 kb interval on chromosome 20 of tetraploid cotton. We identified only one candidate gene with four single nucleotide polymorphisms between parents, among which the single nucleotide polymorphism at the position of 1 082 base pair caused the change of amino acid residue from Arg (3-79) to Lys (T582). The relative expression of the candidate gene in virescent plants was extensively lower than that in normal plants. Nullification of the gene by VIGS significantly turned the green leaf of normal cotton plants into yellow. We named this candidate gene as GhRVL. Conclusions: This study will facilitate the further research on virescent formation, and will be useful for breeding of hybrid cottons.

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Development of Agrobacterium-Mediated Virus-Induced Gene Silencing and Performance Evaluation of Four Marker Genes in Gossypium barbadense

Cited by 71 publications

References 36 publications

The Thioredoxin GbNRX1 Plays a Crucial Role in Homeostasis of Apoplastic Reactive Oxygen Species in Response to Verticillium dahliae Infection in Cotton

The Thioredoxin GbNRX1 Plays a Crucial Role in Homeostasis of Apoplastic Reactive Oxygen Species in Response to Verticillium dahliae Infection in Cotton

The ghr-miR164 and GhNAC100 module participates in cotton plant defence against Verticillium dahliae

Map-based cloning of a recessive gene v1 for virescent leaf expression in cotton (Gossypium spp.)

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