Generation of a transformant library of Colletotrichum gloeosporioides by ATMT and screen of the mutants defective in pathogenicity

Abstract:

[Objective] Generation of C. gloeosporioides T-DNA insertion mutant library through Agrobacterium-mediated transformation was conducted in order to get more insight into the molecular mechanisms underlying the pathogenicity of C. gloeosporioides. [Methods] Agrobacterium containing the binary vector pSULF-gfp was used for transformation of C. gloeosporioides. Chlorimuron-ethyl resistant transformants were screened out and subjected to biological, morphological observation and PCR test. Detached copper-colour rubber leaves were used for pathogenicity assay. [Results] The transformation efficiency was up to 150?400 transformants per 106 conidia and 3721transtormants were generated, 25 transformants defective in pathogenicity were screened out from 3721 transformants. All of the transformants tested remained mitotically stable, maintaining their Chlorimuron-ethyl resistance after tenth generations of growth in the absence of Chlorimuron-ethyl. [Conclution] ATMT can be used to rapidly generate a large library of the fungal transformants, which offers highly efficient means for characterizing the genes that are important for the pathogenicity of C. gloeosporioides and should facilitate further molecular studies of this important plant pathogen.