Using NPG mutant of Aspergillus nidulans which represented a remarkable phenotype at background of wild type allele of velA gene, that is, asexual sporulation was largely inhibited even on a medium with high osmolarity, a genomic DNA fragment in a cosmid vector able to complement a velA1 mutation was isolated. A genetic recombinant with double mutations npgA1 and velA1 that showed normal asexual sporulation was transformed with genomic library DNA constructed in pKBY2 and a transformant showing acondidial phenotype was isolated among about 500 trp+ transformants. The velA1 complementing cosmid DNA was reisolated by in vitro packaging and confirmed by retransformation. The cosmid designated as pVEL1 was able to transform other velA1 mutants at the frequency higher than 80%. The transformants showed several typical velA+ phenotypes including expression of 36Kd protein. The pVEL1 was further subcloned through cotransformation with pKBY2 and 4.7Kb HindIII fragment that retained velA1 complementing activity was finally cloned.