Zymoseptoria tritici is a fungal pathogen of wheat. To decipher molecular interactions between this fungus and its host plant, we developed a strategy for transposon-based insertional mutagenesis. impala is a fungal DNA transposon of the TC1-mariner family moving in host genome by a cut-and-paste mechanism. This transposon is composed of two Terminal Inverted Repeats and a transposase coding gene necessary to excise and reinsert the transposon at a TA site. impala was originally identified in the fungus Fusarium oxysporum, but it transposed successfully in other fungal species. Excision vectors containing an autonomous copy of impala inserted in Aspergillus nidulans nitrate reductase gene were used to select impala excision events in Z. tritici. impala had high rates of excision (85%) and re-insertion (90%) in this fungal species. impala also displayed a positive bias toward genes (90%), preferentially close to transcription starting sites (50%). This pattern was not described before. Overall, impala re-insert randomly in genes from core chromosomes, but seemed to avoid accessory chromosomes poor in expressed genes. We added a strong constitutive promotor (pGdpA) in impala for activation tagging. The chimeric impala:pGpdA transposon was able to excise and re-insert in Z. tritici genome with the same pattern as native impala. We also developed vectors for a double component transposition in which a defective impala carrying pGdpA was trans-activated by an impala transposase encoding gene. These tools provided new methods for mutagenesis in Z. tritici that will be used to identify genes involved in infection. These vectors will be also used to identify mutants of impala transposase with an increased transposition frequency