Fusarium oxysporum f. sp. niveum (FON) race2 isolate was transformed to a hygromycin B (Hyg B)-resistant using two heterologous vectors, pDH25 and CosHyg1. Five and 15 transformed protoplasts, respectively, were obtained per μg of pDH25 and CosHyg1. However, 1-10% of them lost resistance during alternating transfers on Hyg B⁺ and Hyg B^- media. Five pDH25- mediated transformants and twelve CosHygl-mediated transformants were assayed for stability of Hyg B resistance through conidiation. All but two produced Hyg B resistant-microconidia consistently. Of four transformants examined by Southern hybridization, none contained the autonomous replicating vectors. Southern blot analysis showed that multiple integration of vectors occurred. However, no change in pathogenicity was observed among four transformants tested. A genomic library of FL60-3A race 0 isolate was constructed in CosHygl with the average size of 40 kb insert. 960 recombinant clones were hybridized to [³²P]-Iabeled total DNA probe from FL60-3A race 0 and the TX-XID race 2 isolates. Nineteen FL60-3A isolatespecific clones and nine repetitive clones were selected by ifferential hybridization with total DNA probe and used to transform the TX-XID race 2 isolate. Transformation efficiency was 49.4/μg DNA. Nineteen transformants representing 11 specific clones and 7 repetitive clones were bioassayed on watermelon differential cultivars and three transform ants showed the difference in pathogenicity. Probe and in situ digestion analysis of recombinant clone-mediated transformants indicated that all of them were integrative transform ants and had multiple integration of the vectors. They severe rearrangement of vectors after integration due to a large homologous area represented in the insert.