Evaluation of virus-induced gene silencing methods for forage legumes including alfalfa, sainfoin, and fenugreek

2019 ◽  
Vol 99 (6) ◽  
pp. 917-926 ◽  
Author(s):  
Champa Wijekoon ◽  
Stacy D. Singer ◽  
Randall J. Weselake ◽  
Udaya Subedi ◽  
Surya N. Acharya
Keyword(s):  
Gene Silencing ◽  
Plant Species ◽  
Reverse Genetics ◽  
Functional Characterization ◽  
Homologous Gene ◽  
Virus Induced Gene Silencing ◽  
Forage Legumes ◽  
Crop Species ◽  
Barrel Medic ◽  
Trigonella Foenum Graecum L

Virus-induced gene silencing (VIGS) is a rapid reverse genetics tool that has been developed in a wide variety of plant species for assessing gene functions. However, while VIGS has been utilized successfully in the diploid model leguminous species Medicago truncatula (Gaertn.) (barrel medic), such a platform has yet to be established in forage legume crop species. Therefore, we evaluated the effectiveness of this method in forage legumes using a previously developed PEBV (pea early browning virus) system whereby a fragment of the pea (Pisum sativum L.) PHYTOENE DESATURASE (PDS) gene was transferred into a range of alfalfa (Medicago sativa L.), sainfoin (Onobrychis viciifolia Scop.), and fenugreek (Trigonella foenum-graecum L.) cultivars using leaf infiltration and apical meristem injection. Barrel medic was used as a positive control. Gene silencing was observed after 10–15 d through the presence of a leaf bleaching phenotype, and was confirmed using quantitative real-time RT-PCR. Silencing of PDS was achieved in a selection of cultivars in all species assessed, with the highest silencing efficiency apparent in fenugreek. The introduction of a highly homologous gene fragment from a heterologous plant species to target endogenous genes for transient VIGS-based silencing in a range of species of interest represents a potentially useful strategy for the rapid functional characterization of candidate genes in forages.

scholarly journals Virus-induced gene silencing for Asteraceae-a reverse genetics approach for functional genomics in Gerbera hybrida

2012 ◽  
Vol 10 (8) ◽  
pp. 970-978 ◽  
Author(s):  
Xianbao Deng ◽  
Paula Elomaa ◽  
Cuong X. Nguyen ◽  
Timo Hytönen ◽  
Jari P. T. Valkonen ◽  
...  
Keyword(s):  
Gene Silencing ◽  
Functional Genomics ◽  
Reverse Genetics ◽  
Virus Induced Gene Silencing ◽  
Gerbera Hybrida

scholarly journals Virus-induced gene silencing in the perennial woody Paeonia ostii

PeerJ ◽  
2019 ◽  
Vol 7 ◽  
pp. e7001
Author(s):  
Lihang Xie ◽  
Qingyu Zhang ◽  
Daoyang Sun ◽  
Weizong Yang ◽  
Jiayuan Hu ◽  
...  
Keyword(s):  
Gene Silencing ◽  
Fluorescent Protein ◽  
Molecular Genetic ◽  
Functional Characterization ◽  
Tobacco Rattle Virus ◽  
Vacuum Infiltration ◽  
Genomic Research ◽  
Virus Induced Gene Silencing ◽  
Tree Peony ◽  
Medicinal Value

Tree peony is a perennial deciduous shrub with great ornamental and medicinal value. A limitation of its current functional genomic research is the lack of effective molecular genetic tools. Here, the first application of a Tobacco rattle virus (TRV)-based virus-induced gene silencing (VIGS) in the tree peony species Paeonia ostii is presented. Two different approaches, leaf syringe-infiltration and seedling vacuum-infiltration, were utilized for Agrobacterium-mediated inoculation. The vacuum-infiltration was shown to result in a more complete Agrobacterium penetration than syringe-infiltration, and thereby determined as an appropriate inoculation method. The silencing of reporter gene PoPDS encoding phytoene desaturase was achieved in TRV-PoPDS-infected triennial tree peony plantlets, with a typical photobleaching phenotype shown in uppermost newly-sprouted leaves. The endogenous PoPDS transcripts were remarkably down-regulated in VIGS photobleached leaves. Moreover, the green fluorescent protein (GFP) fluorescence was detected in leaves and roots of plants inoculated with TRV-GFP, suggesting the capability of TRV to silence genes in various tissues. Taken together, the data demonstrated that the TRV-based VIGS technique could be adapted for high-throughput functional characterization of genes in tree peony.

scholarly journals Virus-Induced Gene Silencing-Based Functional Characterization of Genes Associated with Powdery Mildew Resistance in Barley

2005 ◽  
Vol 138 (4) ◽  
pp. 2155-2164 ◽  
Author(s):  
Ingo Hein ◽  
Maria Barciszewska-Pacak ◽  
Katarina Hrubikova ◽  
Sandie Williamson ◽  
Malene Dinesen ◽  
...  
Keyword(s):  
Powdery Mildew ◽  
Gene Silencing ◽  
Powdery Mildew Resistance ◽  
Functional Characterization ◽  
Virus Induced Gene Silencing ◽  
Mildew Resistance

Functional characterization of Nicotiana benthamiana homologs of peanut water deficit-induced genes by virus-induced gene silencing

Planta ◽  
2006 ◽  
Vol 225 (3) ◽  
pp. 523-539 ◽  
Author(s):  
M. Senthil-Kumar ◽  
Geetha Govind ◽  
Li Kang ◽  
Kirankumar S. Mysore ◽  
M. Udayakumar
Keyword(s):  
Gene Silencing ◽  
Water Deficit ◽  
Nicotiana Benthamiana ◽  
Functional Characterization ◽  
Virus Induced Gene Silencing ◽  
Induced Genes

Systemic virus-induced gene silencing allows functional characterization of maize genes during biotrophic interaction with Ustilago maydis

2010 ◽  
Vol 189 (2) ◽  
pp. 471-483 ◽  
Author(s):  
Karina van der Linde ◽  
Christine Kastner ◽  
Jochen Kumlehn ◽  
Regine Kahmann ◽  
Gunther Doehlemann
Keyword(s):  
Gene Silencing ◽  
Functional Characterization ◽  
Ustilago Maydis ◽  
Virus Induced Gene Silencing

scholarly journals RNA Interference: A Versatile Tool for Functional Genomics and Unraveling the Genes Required for Viral Disease Resistance in Plants

2019 ◽  
pp. 1-20 ◽  
Author(s):  
Tushar Ranjan ◽  
Namaste Kumari ◽  
Sangita Sahni ◽  
Bishun Deo Prasad
Keyword(s):  
Disease Resistance ◽  
Gene Silencing ◽  
Defense Mechanisms ◽  
Reverse Genetics ◽  
Viral Disease ◽  
Virus Induced Gene Silencing ◽  
Advantages And Disadvantages ◽  
Natural Defense ◽  
Versatile Tool ◽  
Plant Genes

Virus-induced gene silencing (VIGS) is a powerful reverse genetics technology used to unravel the functions of genes. It uses viruses as vectors to carry targeted plant genes. The virus vector is used to induce RNA-mediated silencing of a gene or genes in the host plant. The process of silencing is triggered by dsRNA molecules, the mechanism is explained in this chapter. Over the years a large number of viruses have been modified for use as VIGS vectors and a list of these vectors is also included. As the name suggests, virus-induced gene silencing uses the host plant’s natural defense mechanisms against viral infection to silence plant genes. VIGS is methodologically simple and is widely used to determine gene functions, including disease resistance, abiotic stress, biosynthesis of secondary metabolites and signal transduction pathways. Here, we made an attempt to describe the basic underlying molecular mechanism of VIGS, the methodology and various experimental requirements, as well as its advantages and disadvantages. Finally, we discuss the future prospects of VIGS in relation to CRISPR/Cas9 technology. Besides using it to overexpress or silence genes, VIGS has emerged as the preferred delivery system for the cutting edge CRISPR/Cas9 genome editing technology.

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