Putative transformation of the Alismataceae pathogen, Plectosporium alismatis.

To develop a biocontrol strategy for Alismataceae in rice, Plectosporium alismatis (Oudem.) W.M. Pitt, W. Gams, & U. Braun was putatively transformed with the cutinase gene from Fusarium solani (Mart.) Sacc. f. sp. pisi that is known to be involved in the penetration of plant cuticular structure during fungal invasion. The efficient selection of recombinant transformation events was enhanced by assaying for resistance to hygromycin using the hygromycin B phosphotransferase gene (hph) under the control of Aspergillus nidulans (Eidam) Winter promoter and terminator sequences as a selectable marker. Transformation of fungal protoplasts was achieved by PEG/CaCl 2 treatment with transformation frequencies varying from 6 to 187 transformants per 5 'g of DNA and 2 × 10 6 protoplasts. The antibiotic resistance phenotype appeared to be stable under both selective and nonselective conditions, for at least 4 to 5 generations. Polymerase chain reaction analysis confirmed the presence of vector DNA in transformed cells; however, radioactive probes failed to hybridise to membranes during Southern analysis, and positive confirmation of transformation could not be obtained. Nonintegration of transforming DNA, as opposed to DNA uptake, is believed responsible for episodes of transient transformation indicative of this study.