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

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.

Discovery of a Novel Lineage Burkholderia cepacia ST 1870 Endophytically Isolated from Medicinal Polygala paniculata Which Shows Potent In Vitro Antileishmanial and Antimicrobial Effects

“…Such phytopathogenic fungi and others are responsible for high losses in the production of several crops worldwide [122,123]. C. paradoxa causes the black rot postharvest disease in pineapple [124] and also infects sugarcane [125,126], palm trees, cacao plants, and several other crops [127]. F. verticillioides is a producer of fumonisin, a carcinogenic mycotoxin [128], and other species have been described as emergent and opportunistic pathogens in humans [129].…”

Transferring an Agrobacterium‐mediated transformation protocol across eight genera in the Ceratocystidaceae

“…However, almost all of these were limited to in silico analyses as transformation systems are only available for five Ceratocystidaceae species: three of which are Agrobacterium protocols (including the recent C . albifundus protocol) and the remaining two are PEG/CaCl 2 protoplast transformation systems (Al‐jaaidi, 2007; Loppnau et al, 2004; Niu et al, 2019; Sayari, van der Nest, Steenkamp, Adegeye, et al, 2019; Tzima et al, 2014; Wilson et al, 2020). Considering the importance of this family in plant health, as well as the amount of interest generated through in silico analyses, there is clearly a need to establish more transformation systems that will drive functional studies for these fungi.…”