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Recovery of Mutants Impaired in Pathogenicity After Transposition of Impala in Fusarium oxysporum f. sp. melonis

First author: Dipartimento di Protezione delle Piante, Università di Sassari, Via Enrico De Nicola 9, I-07100 Sassari, Italy; second, fourth, sixth, and seventh authors: Flore Pathogène dans le Sol. INRA-CMSE, UMR BBCE. IPM, 17 rue Sully, BP86510, F.21065 Dijon Cedex, France; and third, fifth, and eighth authors: Institut de Génétique et Microbiologie, Université Paris Sud, Bâtiment 400, F-91405, Orsay, France

The ability of transposon impala to inactivate genes involved in pathogenicity was tested in Fusarium oxysporum f. sp. melonis. Somatic excision of an impala copy inserted in the nitrate reductase-encoding niaD gene was positively selected through a phenotypic assay based on the restoration of nitrate reductase activity. Independent excision events were analyzed molecularly and shown to carry reinsertedimpala in more than 70% of the cases. Mapping of reinserted impala elements on large NotI-restriction fragments showed that impala transposes randomly. By screening 746 revertants on plants, a high proportion (3.5%) of mutants impaired in their pathogenic potential was recovered. According to the kinetics of wilt symptom development, the strains that were impaired in pathogenicity were clustered in three classes: class 1 grouped two strains that never induced Fusarium wilt symptoms on the host plant; class 2 and class 3 grouped 15 and 9 revertants which caused symptoms more than 50 and 30 days after inoculation, respectively. The first results demonstrate the efficiency of transposition in generating mutants affected in pathogenicity, which are usually difficult to obtain by classical mutagenesis, and open the possibility to clone the altered genes with impala as a tag.

Additional keywords: melon, transposon tagging.