Filamentous fungi are indispensable biotechnological tools for the production of organic chemicals, enzymes, and antibiotics. Most of the strains used for industrial applications have been--and still are--screened and improved by classical mutagenesis. Sexual crossing approaches would yield considerable advantages for research and industrial strain improvement, but interestingly, industrially applied filamentous fungal species have so far been considered to be largely asexual. This is also true for the ascomycete Trichoderma reesei (anamorph of Hypocrea jecorina), which is used for production of cellulolytic and hemicellulolytic enzymes. In this study, we report that T. reesei QM6a has a MAT1-2 mating type locus, and the identification of its respective mating type counterpart, MAT1-1, in natural isolates of H. jecorina, thus proving that this is a heterothallic species. After being considered asexual since its discovery more than 50 years ago, we were now able to induce sexual reproduction of T. reesei QM6a and obtained fertilized stromata and mature ascospores. This sexual crossing approach therefore opens up perspectives for biotechnologically important fungi. Our findings provide a tool for fast and efficient industrial strain improvement in T. reesei, thus boosting research toward economically feasible biofuel production. In addition, knowledge of MAT-loci and sexual crossing techniques will facilitate research with other Trichoderma spp. relevant for agriculture and human health.