Vitis rotundifolia (Muscadine grapes), a native species characterized with multiple resistance to grape diseases and insects, are cultivated throughout the southeastern U.S. for fresh fruit and processing. However, the species falls short of consumer's expectation as fresh fruit due to its seediness and thick skin. However, Vitis vinifera, a predominant Vitis species grown worldwide possesses good fruit characteristics such as seedlessness and edible skin but is susceptible to many diseases. Attempts to produce rotundifolia-vinifera hybrids to combine good fruit quality and disease resistance of both into F1 hybrids have been made by grape breeders for many years. Limited success was only reported when the V. vinifera was used as seed parents. Pollinating seedless vinifera pollen onV. rotundifolia stigma was made in 1993 and 1994. More than 20,000 flowers from 34 cross combinations were pollinated. These crosses were made to see if there is any chance to produce hybrids when muscadine grapes were used as female parent and specifically to introgress the seedlessness from European grapes into muscadine grapes. A few hundred seeds were collected from these crosses and germinated in a greenhouse. Two seedlings were clearly distinguished from the others with morphology intermediate between muscadine and the vinifera grapes, while the rest looked straight muscadine grapes derived from possible contaminated pollination. This conclusion was further confirmed by isozyme and DNA markers. One of the seedlings produced from the cross of `Jumbo' × `Thompson Seedless' grew vigorously and has been setting fruit since 1996. Fruit are mixture of stenospermocarpic and pathonocarpic seedlessness. Fruit setting and pollen viability test indicated that this hybrid is at least partly self-fertile. Many other characteristics of the hybrid, such as leaves, stems, tendrils, time of budbreak, bloom date, and ripen date are intermediate between muscadine and bunch grapes. The hybrid is resistant to Pierce's disease, anthracnose disease, and downy mildew, which are the limited factor to growing V. vinifera in the hot and humid southeastern U.S. This is the first report of a seedless hybrid from V. rotundifolia × V. vinifera.