The pOT and pLOB vector systems: Improving ease of transgene expression in Botrytis cinerea

Patel, Risha M.; Heneghan, Mary N.; Kan, Jan A.L. van; Bailey, Andy M.; Foster, Gary D.

doi:10.2323/jgam.54.367

Cited by 21 publications

(14 citation statements)

References 41 publications

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“…Thus, the 5′- and 3′-non-coding regions of bcwcl1 were amplified from genomic DNA of B. cinerea B05.10 using the primer pairs indicated in Table S2. The hygromycin ( hph ) resistance cassette was amplified from vector pLOB1 [51] (Δ bcwcl1 , mutant 1, replacement cassette A) or pCSN44 (obtained from the Fungal Genetics Stock Center, [52]) as template (Δ bcwcl1 , mutant 2 and 3; replacement cassette B). Primers employed for these PCR reactions contained 30-bp-overlapping regions, thus allowing homologous recombination.…”

Section: Methodsmentioning

confidence: 99%

Assessing the Effects of Light on Differentiation and Virulence of the Plant Pathogen Botrytis cinerea: Characterization of the White Collar Complex

et al. 2013

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Organisms are exposed to a tough environment, where acute daily challenges, like light, can strongly affect several aspects of an individual's physiology, including pathogenesis. While several fungal models have been widely employed to understand the physiological and molecular events associated with light perception, various other agricultural-relevant fungi still remain, in terms of their responsiveness to light, in the dark. The fungus Botrytis cinerea is an aggressive pathogen able to cause disease on a wide range of plant species. Natural B. cinerea isolates exhibit a high degree of diversity in their predominant mode of reproduction. Thus, the majority of naturally occurring strains are known to reproduce asexually via conidia and sclerotia, and sexually via apothecia. Studies from the 1970′s reported on specific developmental responses to treatments with near-UV, blue, red and far-red light. To unravel the signaling machinery triggering development – and possibly also connected with virulence – we initiated the functional characterization of the transcription factor/photoreceptor BcWCL1 and its partner BcWCL2, that form the White Collar Complex (WCC) in B. cinerea. Using mutants either abolished in or exhibiting enhanced WCC signaling (overexpression of both bcwcl1 and bcwcl2), we demonstrate that the WCC is an integral part of the mentioned machinery by mediating transcriptional responses to white light and the inhibition of conidiation in response to this stimulus. Furthermore, the WCC is required for coping with excessive light, oxidative stress and also to achieve full virulence. Although several transcriptional responses are abolished in the absence of bcwcl1, the expression of some genes is still light induced and a distinct conidiation pattern in response to daily light oscillations is enhanced, revealing a complex underlying photobiology. Though overlaps with well-studied fungal systems exist, the light-associated machinery of B. cinerea appears more complex than those of Neurospora crassa and Aspergillus nidulans.

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Section: Methodsmentioning

confidence: 99%

Assessing the Effects of Light on Differentiation and Virulence of the Plant Pathogen Botrytis cinerea: Characterization of the White Collar Complex

et al. 2013

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“…Vector pLOB7 (Patel et al, 2008) was used to construct the replacement vector. A replacement vector for rho3 was designed as described in Fig.…”

Section: Vector Constructionsmentioning

confidence: 99%

Function of small GTPase Rho3 in regulating growth, conidiation and virulence of Botrytis cinerea

Qin

et al. 2015

Fungal Genetics and Biology

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“…Putative transformants were screened for DsRed expression in situ and following subculture onto fresh selection plates using a DM-LB Leica microscope fitted with a red-shifted TRITC filter (excitation 545/30 nm, emission 620/60 nm) [18], [19].…”

Section: Methodsmentioning

confidence: 99%

Investigating Agrobacterium-Mediated Transformation of Verticillium albo-atrum on Plant Surfaces

2010

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Background Agrobacterium tumefaciens has long been known to transform plant tissue in nature as part of its infection process. This natural mechanism has been utilised over the last few decades in laboratories world wide to genetically manipulate many species of plants. More recently this technology has been successfully applied to non-plant organisms in the laboratory, including fungi, where the plant wound hormone acetosyringone, an inducer of transformation, is supplied exogenously. In the natural environment it is possible that Agrobacterium and fungi may encounter each other at plant wound sites, where acetosyringone would be present, raising the possibility of natural gene transfer from bacterium to fungus.Methodology/Principal FindingsWe investigate this hypothesis through the development of experiments designed to replicate such a situation at a plant wound site. A. tumefaciens harbouring the plasmid pCAMDsRed was co-cultivated with the common plant pathogenic fungus Verticillium albo-atrum on a range of wounded plant tissues. Fungal transformants were obtained from co-cultivation on a range of plant tissue types, demonstrating that plant tissue provides sufficient vir gene inducers to allow A. tumefaciens to transform fungi in planta.Conclusions/SignificanceThis work raises interesting questions about whether A. tumefaciens may be able to transform organisms other than plants in nature, or indeed should be considered during GM risk assessments, with further investigations required to determine whether this phenomenon has already occurred in nature.

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The pOT and pLOB vector systems: Improving ease of transgene expression in Botrytis cinerea

Cited by 21 publications

References 41 publications

Assessing the Effects of Light on Differentiation and Virulence of the Plant Pathogen Botrytis cinerea: Characterization of the White Collar Complex

Assessing the Effects of Light on Differentiation and Virulence of the Plant Pathogen Botrytis cinerea: Characterization of the White Collar Complex

Function of small GTPase Rho3 in regulating growth, conidiation and virulence of Botrytis cinerea

Investigating Agrobacterium-Mediated Transformation of Verticillium albo-atrum on Plant Surfaces

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