Agrobacterium tumefaciens-mediated genetic transformation of the entomopathogenic fungus Beauveria bassiana.
Agrobacterium tumefaciens-mediated transformation (agro-transformation) was successfully applied to the entomogenous fungus Beauveria bassiana. Conidia of B. bassiana were transformed to hygromycin B resistance using the hph gene of Escherichia coli as the selective trait, under the control of a heterologous fungal promoter and the Aspergillus nidulans trpC terminator. The efficiency of transformation was up to 28 and 96 transformants per 10(4) and 10(5) target conidia, respectively, using three distinct vectors. High mitotic stability of the transformants (80-100%) was demonstrated after five successive transfers on a nonselective medium. Abortive transformants were observed for all the hph(r) vectors used. Putative transformants were analysed for the presence of the hph gene by PCR and Southern analysis. The latter analysis revealed the integration of two or more copies of the hph gene in the genome. The agro-transformation method was found to be effective for the isolation of B. bassiana hygromycin resistant transformants and may represent a useful tool for insertional mutagenesis studies in this fungus.
dos Reis
Maria Cecília
MC
Departamento de Microbiologia, Universidade Estadual de Londrina, P.O. Box 6001, Londrina 86051-990, Brazil.
Pelegrinelli Fungaro
Maria Helena
MH
Delgado Duarte
Rubens Tadeu
RT
Furlaneto
Luciana
L
Furlaneto
Marcia Cristina
MC
D016960:Agrobacterium tumefaciens / Q000235:genetics*; D000818:Animals; D024881:Drug Resistance, Bacterial / Q000235:genetics; D004926:Escherichia coli / Q000187:drug effects / Q000235:genetics; D005798:Genes, Bacterial; D006921:Hygromycin B / Q000494:pharmacology; D006999:Hypocreales / Q000235:genetics* / Q000472:pathogenicity*; D007313:Insecta / Q000382:microbiology; D008828:Microbiological Techniques; D010572:Pest Control, Biological; D014170:Transformation, Genetic
D006921:Hygromycin B