A single clone harboring tRNA^(AsP) gene was isolated by colony hybridization to the total tRNA gene clone of Aspergillus nidulans using the previously sequenced tRNA^(AsP)probe. The gene was localized and sequenced by the method of Maxam and Gilbert. The sequence analysis revealed that the tRNA^(AsP) gene has the anticodon of GUC, and contains an intervening sequence of 15 bases. The nucleotide sequence of this tRNA^(AsP) gene was compared with tRNA^(AsP`)s of the other organisms. From this, some structural features in the flanking sequences and the gene were discussed.