Abstract: In this study, we developed an Agrobacterium tumefaciensmediated transformation system for Pestalotiopsis heterocornis by using A. tumefaciens strain AGL1 which carried plasmid pBHt1 harboring the hygromycin B phosphotransferase gene(hph). The results showed a total of 200-300 transformants were obtained per 1×106 spores when the spore suspension were cocultivated with A. tumefaciens cells,the optimal conditions were OD600=0.3, 25 ℃,and 48 h cultivation in the presence of induction medium containing acetosyringone at 200 μg/mL. Selection medium performed best with concentrations of 250 μg/mL of hygromycin B and 250 μg/mL of cefotaxime sodium in PDA. 〖JP2〗To detect the heredity stability, ten random transformants were cultivated on the PDA medium for five generations, colony PCR of each generation obtained expected amplification bands, which showed that the transformants exhibit high heredity stability. The transformation system will be useful for further studies of functional genes and hostpathogen interaction in P. heterocornis.