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

1
Key Laboratory of Biopesticide and Chemical Biology, Ministry of Education, Fujian Agriculture and Forestry University, Fuzhou 350002, China
2
Coconut Research Institute, Chinese Academy of Tropical Agricultural Sciences, Wenchang 571339, China
*
Author to whom correspondence should be addressed.
Int. J. Mol. Sci. 2019, 20(10), 2387; https://doi.org/10.3390/ijms20102387
Received: 10 April 2019 / Revised: 11 May 2019 / Accepted: 11 May 2019 / Published: 14 May 2019
(This article belongs to the Section Molecular Microbiology)
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Abstract

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. View Full-Text
Keywords: stem-bleeding disease of coconut; Ceratocystis paradoxa; gfp gene; transformation; protoplast stem-bleeding disease of coconut; Ceratocystis paradoxa; gfp gene; transformation; protoplast
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Niu, X.; Pei, M.; Liang, C.; Lv, Y.; Wu, X.; Zhang, R.; Lu, G.; Yu, F.; Zhu, H.; Qin, W. Genetic Transformation and Green Fluorescent Protein Labeling in Ceratocystis paradoxa from Coconut. Int. J. Mol. Sci. 2019, 20, 2387.

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