Filamentous fungi play an important role in industry, agriculture, medicine and biological fundamental research. More and more attention has been paid to fungal breeding and gene functional analysis via genetic transformation technology. However, the extraction of fungal DNA was labor intensive and time-consuming, which makes it difficult to perform high-throughput screening of transformants through PCR technology. In this paper, a rapid DNA extraction method was established for filamentous fungi (take Aspergillus for example). DNA was easily obtained by exposing the fungal mycelia in 10 × TE buffer to microwave. RAPD and PCR experiments demonstrated that the extracted DNA satisfied the requirements of PCR amplification. These results provided a foundation for the high-throughput screening of filamentous fungal transformants.