Genetic Transformation and Green Fluorescent Protein Labeling in Ceratocystis paradoxa from Coconut

Ceratocystis paradoxa, the causal agent of stem-bleeding disease of the coconut palm, causes great losses to the global coconut industry. As the mechanism of pathogenicity of C. paradoxa has not been determined, an exogenous gene marker was introduced into the fungus. In this study, pCT74-sGFP, which contains the green fluorescent protein (GFP) gene, and the hygromycin B resistance gene as a selective marker, was used as an expression vector. Several protoplast release buffers were compared to optimize protoplast preparation. The plasmid pCT74-sGFP was successfully transformed into the genome of C. paradoxa, which was verified using polymerase chain reaction and green fluorescence detection. The transformants did not exhibit any obvious differences from the wild-type isolates in terms of growth and morphological characteristics. Pathogenicity tests showed that the transformation process did not alter the virulence of the X-3314 C. paradoxa strain. This is the first report on the polyethylene glycol-mediated transformation of C. paradoxa carrying a ‘reporter’ gene GFP that was stably and efficiently expressed in the transformants. These findings provide a basis for future functional genomics studies of C. paradoxa and offer a novel opportunity to track the infection process of C. paradoxa.

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Interaction of the Mite Aceria mangiferae with Fusarium mangiferae, the Causal Agent of Mango Malformation Disease

Phytopathology ◽

10.1094/phyto-99-2-0152 ◽

2009 ◽

Vol 99 (2) ◽

pp. 152-159 ◽

Cited By ~ 14

Author(s):

E. Gamliel-Atinsky ◽

S. Freeman ◽

A. Sztejnberg ◽

M. Maymon ◽

R. Ochoa ◽

...

Keyword(s):

Green Fluorescent Protein ◽

Fungal Infection ◽

Fluorescent Protein ◽

Infection Process ◽

Apical Bud ◽

Apical Buds ◽

Green Fluorescent ◽

Mango Malformation ◽

Fusarium Mangiferae

The role of the mango bud mite, Aceria mangiferae, in carrying conidia of Fusarium mangiferae, vectoring them into potential infection sites, and assisting fungal infection and dissemination was studied. Following the mite's exposure to a green fluorescent protein-marked isolate, conidia were observed clinging to the mite's body. Agar plugs bearing either bud mites or the pathogen were placed on leaves near the apical buds of potted mango plants. Conidia were found in bud bracts only when both mites and conidia were co-inoculated on the plant, demonstrating that the mite vectored the conidia into the apical bud. Potted mango plants were inoculated with conidia in the presence or absence of mites. Frequency and severity of infected buds were significantly higher in the presence of mites, revealing their significant role in the fungal infection process. Conidia and mite presence were monitored with traps in a diseased orchard over a 2-year period. No windborne bud mites bearing conidia were found; however, high numbers of windborne conidia were detected in the traps. These results suggest that A. mangiferae can carry and vector conidia between buds and assist in fungal penetration but does not play a role in the aerial dissemination of conidia between trees.

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Studies on Vascular Infection of Fusarium oxysporum f. sp. cubense Race 4 in Banana by Field Survey and Green Fluorescent Protein Reporter

International Journal of Phytopathology ◽

10.33687/phytopath.002.01.0064 ◽

2013 ◽

Vol 2 (1) ◽

pp. 44-51 ◽

Cited By ~ 1

Author(s):

Rong F. Xiao ◽

Yu-Jing Zhu ◽

Yan-Dan Li ◽

Bo Liu

Keyword(s):

Green Fluorescent Protein ◽

Fusarium Oxysporum ◽

Field Survey ◽

Fluorescent Protein ◽

Infection Process ◽

Gene Encoding ◽

Effective Prevention ◽

Green Fluorescent ◽

Banana Wilt ◽

Race 4

Fusarium wilt of banana (Musa spp.) caused by Fusarium oxysporum f. sp. cubense (Foc) is one of the most serious banana fungal diseases in the world. Understanding the infection process of Foc is important for development of effective ways in disease control. In order to follow infection and colonization of this pathogen from root to rhizome and pseudostem tissues of banana, a highly pathogenic strain FJAT-3076 of Foc race 4 (Foc4) was transformed with gene encoding green fluorescent protein (GFP) and the fungus carrying gfp (FJAT-3076-GFP) was used to inoculate banana plants (Cavendish cv. B.F.). After inoculation for 3 to 10 d, it was observed that the conidia and their germ-tubes had penetrated into epidermis of young roots. The hyphae were found inside the root xylem 10 d after inoculation in the rhizome and pseudostem xylem after inoculation for 17 d. All plants infected by Foc died in 24 d after inoculation. It was also observed that Foc had spread all over the xylem and part of hyphae reached the pseudostem surface. Hyphal population was found the highest in the pseudostem, lower in root and least in rhizome. Field survey confirmed that Foc4 were mostly present in the base of pseudostem and less in the rhizome. Thus, effective prevention of the Foc hyphae movement from the rhizome up to the pseudostem might delay or control banana wilt disease.

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