One Step Construction of Agrobacterium-Recombination-ready-plasmids (OSCAR), an efficient and robust tool for ATMT based gene deletion construction in fungi

Paz, Zahi; García-Pedrajas, Maria D.; Andrews, Davıd L.; Klosterman, Steven J.; Baeza-Montañez, Lourdes; Gold, Scott E.

doi:10.1016/j.fgb.2011.02.003

Cited by 83 publications

(85 citation statements)

References 33 publications

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“…Reviewing the literature, we found that ATMT has been widely used as a method to identify the virulence factors in different fungal species (Li et al, 2012;Paz et al, 2011;Wang et al, 2016;Zhang et al, 2015). In the present study, Table 1.…”

Section: Discussionmentioning

confidence: 51%

“…ATMT has been established as an important technique to acquire insertional mutant libraries and to identify pathogenicity-related genes. Furthermore, the high efficiency of targeted gene knock-out also permits quick functional analysis of putative pathogenicity-related genes (Li et al, 2012;Paz et al, 2011;Wang et al, 2016). In this study, the characterization of the C2M52 mutant was carried out to show the insertion of T-DNA in the promoter region of the hypothetical protein (BcHP) gene.…”

Section: Introductionmentioning

confidence: 99%

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Identification of new pathogenicity related to the gene encoding hypothetical protein in the gray mold fungus (Botrytis cinerea)

DafaAlla¹,

Abdalla²,

Algaili³

et al. 2017

Aust J Crop Sci

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Botrytis cinerea is an important necrotrophic fungus that causes gray mold disease in various important crop plants, leading to substantial economic losses. Herein, Agrobacterium tumefaciens-mediated transformation (ATMT) technique was used to create an insertional mutant library of B. cinerea to investigate its pathogenicity-related genes. Among 1,734 transformations only 46 mutants decreased the pathogenicity on tomato leaves. C2M52 is one of the 46 mutants that its pathogenicity reduction was analyzed deeply. The T-DNA integrated into the promoter region of the hypothetical protein (BcHP) gene. The gene replacement approach via homologous recombination was used to generate knock-out mutant (△bchp) and its functional complementary (△bchp-C) strains. The results warranted that the △bchp mutant strain displayed a significant difference in the virulence on tomato leaves relative to the wild type. These results were consistent with the attenuated pathogenicity caused by T-DNA integration mutant (C2M52). That subsequently revealed the crucial role of BcHP gene in the pathogenicity of B. cinerea.

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Section: Discussionmentioning

confidence: 51%

Section: Introductionmentioning

confidence: 99%

Identification of new pathogenicity related to the gene encoding hypothetical protein in the gray mold fungus (Botrytis cinerea)

DafaAlla¹,

Abdalla²,

Algaili³

et al. 2017

Aust J Crop Sci

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show abstract

“…For FUBT disruption, the one-step construction of Agrobacterium-recombinationready-plasmids (OSCAR) method was used to construct the transformation plasmids utilizing a Gateway cloning technique (Paz et al, 2011). The resulting plasmid (pOSCAR-FATR) was transformed into AGL-1.…”

Section: Methodsmentioning

confidence: 99%

FUBT, a putative MFS transporter, promotes secretion of fusaric acid in the cotton pathogen Fusarium oxysporum f. sp. vasinfectum

Crutcher¹,

Liu²,

Puckhaber³

et al. 2015

Microbiology

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Fusaric acid (FA) is a key component in virulence and symptom development in cotton during infection by Fusarium oxysporum. A putative major facilitator superfamily (MFS) transporter gene was identified downstream of the polyketide synthase gene responsible for the biosynthesis of FA in a region previously believed to be unrelated to the known FA gene cluster. Disruption of the transporter gene, designated FUBT, resulted in loss of FA secretion, decrease in FA production and a decrease in resistance to high concentrations of FA. Uptake of exogenous FA was unaffected in the disruption transformants, suggesting that FA enters the cell in Fusarium by an independent mechanism. Thus, FUBT is involved both in the extracellular transport of FA and in resistance of F. oxysporum to this non-specific toxin. A potential secondary resistance mechanism, the production of FA derivatives, was observed in FUBT deletion mutants. Molecular analysis of key biochemical processes in the production of FA could lead to future host plant resistance to Fusarium pathogens.

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“…Generating constructs using traditional approaches like digestion/ligation is labour intensive, time-consuming, relatively expensive as it requires several cloning steps. Hence, recently several studies have been conducted to improve or develop new genetic tools for large-scale functional analyses (Paz et al, 2011;Shafran et al, 2008;Zhu et al, 2009). Among these, the Gateway Ò cloning technology has attracted molecular biologists from different disciplines due to its amenability and robustness (Schoberle et al, 2013).…”

Section: Introductionmentioning

confidence: 99%

“…Among these, the Gateway Ò cloning technology has attracted molecular biologists from different disciplines due to its amenability and robustness (Schoberle et al, 2013). To date, a few methods or constructs have been developed using this technology for the functional analyses of genes in plant pathogenic fungi (Abe et al, 2006;Nakagawa et al, 2007;Paz et al, 2011;Shafran et al, 2008;Zhu et al, 2009). For instance, the One Step Construction of Agrobacterium-Recombination-ready-plasmids (OSCAR) has been developed to create deletion constructs for Agrobacterium tumefaciens mediated transformation (Paz et al, 2011).…”

Section: Introductionmentioning

confidence: 99%

Flexible gateway constructs for functional analyses of genes in plant pathogenic fungi

Mehrabi

Gohari

Silva

et al. 2015

Fungal Genetics and Biology

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One Step Construction of Agrobacterium-Recombination-ready-plasmids (OSCAR), an efficient and robust tool for ATMT based gene deletion construction in fungi

Cited by 83 publications

References 33 publications

Identification of new pathogenicity related to the gene encoding hypothetical protein in the gray mold fungus (Botrytis cinerea)

Identification of new pathogenicity related to the gene encoding hypothetical protein in the gray mold fungus (Botrytis cinerea)

FUBT, a putative MFS transporter, promotes secretion of fusaric acid in the cotton pathogen Fusarium oxysporum f. sp. vasinfectum

Flexible gateway constructs for functional analyses of genes in plant pathogenic fungi

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