The parents and progeny from two crosses (Cayuga White x Aurore and NY62.136.2 x Yates) were examined for the presence of DNA restriction fragment length polymorphisms (RFLPs). Seventeen independent DNA sequences were used in the analysis 15 obtained from a grape Pstl genomic library and two heterologous probes obtained from other laboratories. Most of the low copy cloned sequences hybridized to more than two restriction fragments possibly reflecting the polyploid nature of the Vitis genome. Nine of the probes detected RFLPs between parents. Analysis of the progenies (F1generation) revealed segregation for nine distinct polymorphisms generated by seven of the probes.Thus a relatively high level of polymorphism among parents as well as heterozygosity within each parent was evident. Most RFLPs gave segregation ratios close to the 1: 1 ratio predicted for a locus heterozygous in one parent. However four differences between parental phenotypes did not segregated in the progeny and in three instances fragments present in both parents segregated in the progeny. These peculiar results may be explained by accounting for heterozygosity or homozygosity respectively for the DNA segment that generates the polymorphism. We conclude that RFLP studies can be performed on the first filial generation in woody perennials such as Vitis that have a relatively high level of heterozygosity in the genome.