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首页> 外文期刊>Australasian Plant Pathology >Transformation of Fusarium oxysporum f. sp cubense, causal agent of Fusarium wilt of banana, with the green fluorescent protein (GFP) gene
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Transformation of Fusarium oxysporum f. sp cubense, causal agent of Fusarium wilt of banana, with the green fluorescent protein (GFP) gene

机译：尖孢镰刀菌的转化香蕉枯萎病病原菌sp cubense，具有绿色荧光蛋白（GFP）基因

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摘要

Fusarium oxysporum f. sp. cubense (Foc) is the causal agent of Fusarium wilt (Panama disease) of bananas in most tropical and subtropical banana-producing regions of the world. The fungus infects through roots, colonises the rhizomes and eventually blocks the vascular system of the pseudostems, resulting in plant death. The green fluorescent protein (GFP) emits green fluorescence when excited by blue light, making it a useful tool to study early stages of fungal infection. The objective of this study was to transform Foc isolates with the GFP gene. Isolates representing 'subtropical' race 4 of the fungus were transformed with the sGFP derivative using hygromycin as a selectable marker. Efficiency and transformation of spheroplasts depended on mycelium age, the choice of enzymes and the temperature and duration of incubation. The transformed isolates did not differ markedly from the wild type isolates in growth and morphological characteristics in vitro. Fluorescence microscopy showed expression of the green fluorescent protein in fungal structures. The presence of the GFP DNA in the fungal cells was confirmed by PCR using a GFP-specific primer pair and Southern blot analysis. Pathogenicity tests showed that the transformation process did not alter pathogenicity of Foc isolates. Fungal hyphae within tissues of infected plants could be seen to fluoresce and the transformed fungus was re-isolated from artificially inoculated plants. Transformants of Foc will facilitate future infection studies with this pathogen on banana.
机译：尖孢镰刀菌f。 sp。在世界上大多数热带和亚热带香蕉产区，cubanse（Foc）是香蕉枯萎病（巴拿马病）的病原体。真菌通过根部感染，定居在根茎上，并最终阻塞假茎的血管系统，导致植物死亡。当被蓝光激发时，绿色荧光蛋白（GFP）发出绿色荧光，使其成为研究真菌感染早期的有用工具。这项研究的目的是用GFP基因转化Foc分离株。使用潮霉素作为选择标记，用sGFP衍生物转化代表真菌“亚热带”第4类的菌株。原生质球的效率和转化取决于菌丝体年龄，酶的选择以及孵育的温度和持续时间。在体外，转化的分离株与野生型分离株在生长和形态学特征上没有显着差异。荧光显微镜显示绿色荧光蛋白在真菌结构中的表达。通过PCR使用GFP特异性引物对和Southern印迹分析确认了真菌细胞中GFP DNA的存在。致病性测试表明，转化过程并未改变Foc分离株的致病性。可以看到被感染植物组织内的真菌菌丝发荧光，并且从人工接种的植物中重新分离了转化的真菌。 Foc的转化子将促进该病原体在香蕉上的进一步感染研究。

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