scholarly journals Simple and efficient heterologous expression of necrosis‐inducing effectors using the model plant Nicotiana benthamiana

Plant Direct ◽  
2021 ◽  
Vol 5 (8) ◽  
Author(s):  
Bayantes Dagvadorj ◽  
Peter S. Solomon
Keyword(s):  
Heterologous Expression ◽  
Nicotiana Benthamiana ◽  
Model Plant

scholarly journals Simple and efficient heterologous expression of necrosis-inducing effectors using the model plant Nicotiana benthamiana

2021 ◽  
Author(s):  
Bayantes Dagvadorj ◽  
Peter Solomon
Keyword(s):  
Heterologous Expression ◽  
High Throughput ◽  
Posttranslational Modifications ◽  
Nicotiana Benthamiana ◽  
Fungal Pathogens ◽  
Host Plants ◽  
Plant Pathogens ◽  
Expression Systems ◽  
In Planta ◽  
Model Plant

Plant fungal pathogens cause devastating diseases on cereal plants and threaten global food security. During infection, these pathogens secrete proteinaceous effectors that promote disease. Some of these effectors from necrotrophic plant pathogens induce a cell death response (necrosis), which facilitates pathogen growth in planta. Characterisation of these effectors typically requires heterologous expression and microbial expression systems such as bacteria and yeast are the predominantly used. However, microbial expression systems often require optimization for any given effector and are, in general, not suitable for effectors involving cysteine bridges and posttranslational modifications for activity. Here, we describe a simple and efficient method for expressing such effectors in the model plant Nicotiana benthamiana. Briefly, an effector protein is transiently expressed and secreted into the apoplast of N. benthamiana by Agrobacterium-mediated infiltration. Two-to-three days subsequent to agroinfiltration, the apoplast from the infiltrated leaves is extracted and can be directly used for phenotyping on host plants. The efficacy of this approach was demonstrated by expressing the ToxA, Tox3 and Tox1 necrosis-inducing effectors from Parastagonospora nodorum. All three effectors produced in N. benthamiana were capable of inducing necrosis in wheat lines, and two of three showed visible bands on Coomassie-stained gel. These data suggest that N. benthamiana-agroinfiltration system is a feasible tool to obtain fungal effectors, especially those that require disulfide bonds and posttranslational modifications. Furthermore, due to the low number of proteins typically observed in the apoplast (compared to intracellular), this simple and high-throughput approach circumvents the requirement to lyse cells and further purify the target proteins that is required in other heterologous systems. Because of its simplicity and potential for high-throughput, this method is highly amenable to the phenotyping of candidate protein effectors on host plants.

scholarly journals Heterologous Expression Screens in Nicotiana benthamiana Identify a Candidate Effector of the Wheat Yellow Rust Pathogen that Associates with Processing Bodies

PLoS ONE ◽  
2016 ◽  
Vol 11 (2) ◽  
pp. e0149035 ◽  
Author(s):  
Benjamin Petre ◽  
Diane G. O. Saunders ◽  
Jan Sklenar ◽  
Cécile Lorrain ◽  
Ksenia V. Krasileva ◽  
...  
Keyword(s):  
Heterologous Expression ◽  
Nicotiana Benthamiana ◽  
Yellow Rust ◽  
Processing Bodies ◽  
Rust Pathogen

scholarly journals A Pseudomonas syringae pv. tomato DC3000 mutant lacking the type III effector HopQ1-1 is able to cause disease in the model plant Nicotiana benthamiana

2007 ◽  
Vol 51 (1) ◽  
pp. 32-46 ◽  
Author(s):  
Chia-Fong Wei ◽  
Brian H. Kvitko ◽  
Rena Shimizu ◽  
Emerson Crabill ◽  
James R. Alfano ◽  
...  
Keyword(s):  
Pseudomonas Syringae ◽  
Nicotiana Benthamiana ◽  
Tomato Dc3000 ◽  
Type Iii Effector ◽  
Model Plant ◽  
Type Iii

scholarly journals Heterologous expression of the isopimaric acid pathway in Nicotiana benthamiana and the effect of N-terminal modifications of the involved cytochrome P450 enzyme

2015 ◽  
Vol 9 (1) ◽  
Author(s):  
Thiyagarajan Gnanasekaran ◽  
Konstantinos Vavitsas ◽  
Johan Andersen-Ranberg ◽  
Agnieszka Zygadlo Nielsen ◽  
Carl Erik Olsen ◽  
...  
Keyword(s):  
Cytochrome P450 ◽  
Heterologous Expression ◽  
Nicotiana Benthamiana ◽  
Cytochrome P450 Enzyme ◽  
Isopimaric Acid ◽  
P450 Enzyme ◽  
Acid Pathway

scholarly journals Reducing biomass recalcitrance by heterologous expression of a bacterial peroxidase in tobacco (Nicotiana benthamiana)

2017 ◽  
Vol 7 (1) ◽  
Author(s):  
Ayalew Ligaba-Osena ◽  
Bertrand Hankoua ◽  
Kay DiMarco ◽  
Robert Pace ◽  
Mark Crocker ◽  
...  
Keyword(s):  
Heterologous Expression ◽  
Nicotiana Benthamiana ◽  
Biomass Recalcitrance

Micro RNA ‐induced gene silencing strategy for the delivery of siRNAs targeting Meloidogyne incognita in a model plant Nicotiana benthamiana

2021 ◽  
Author(s):  
Alkesh Hada ◽  
Basavaprabhu L. Patil ◽  
Akansha Bajpai ◽  
Karthik Kesiraju ◽  
Savithramma Dinesh‐Kumar ◽  
...  
Keyword(s):  
Gene Silencing ◽  
Meloidogyne Incognita ◽  
Nicotiana Benthamiana ◽  
Micro Rna ◽  
Model Plant

scholarly journals Validating Plant Genes Involved in Pepper Yellow Leaf Curl Indonesia Virus Infection Using VIGS in Model Plant Nicotiana benthamiana

2020 ◽  
Vol 16 (1) ◽  
pp. 7
Author(s):  
Kusumawaty Kusumanegara ◽  
Masanori Kaido ◽  
Kazuyuki Mise
Keyword(s):  
Viral Infection ◽  
Nicotiana Benthamiana ◽  
Tobacco Rattle Virus ◽  
Chili Pepper ◽  
Resistant Variety ◽  
Post Inoculation ◽  
Yellow Leaf ◽  
Model Plant ◽  
Plant Genes ◽  
Leaf Curl

<p>Pepper yellow leaf curl disease caused by Pepper yellow leaf curl Indonesia virus (PepYLCIV) has become a challenge to chili pepper cultivation. Development of resistant variety by utilizing recessive resistance gene is expected to control the disease in the field. This study aimed to validate three plant genes, namely deltaCOP, hsc70, and BAM1, in PepYLCIV infection by applying Virus-induced Gene Silencing (VIGS) in a model plant, wild type Nicotiana benthamiana. PepYLCIV and construct of Tobacco rattle virus (TRV) which induced silencing of each gene were co-inoculated into N. benthamiana plants through agroinfiltration. Gene expression and the relative amount of viral DNA were determined by quantitative reverse transcription PCR (qRT-PCR) and quantitative PCR (qPCR), respectively, at 15 days post inoculation. The results showed a decreased level of deltaCOP, hsc70, and BAM1 expressions to 66.4%, 53.0%, and 47.0%, respectively, compared to that in the control (100%). Silencing of the three genes decreased the accumulation of PepYLCIV to 0.1%, 18.4%, and 63.0%, respectively, compared to that in the control. deltaCOP and hsc70 genes were indicated to be involved in the viral infection and could be good candidate genes for obtaining chili pepper varieties resistant to PepYLCIV. This result affirmed that the reverse genetics technique could be an alternative approach for identifying plant genes involved in viral infection, including PepYLCIV. The use of an infectious clone in this study allows the virus inoculations could be carried out without rearing and maintaining its natural vector, hence reduces the risk of virus transmission to healthy plants.</p>

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