We have identified and analyzed a meiotic reciprocal recombination hot spot in S. cerevisiae. We find that double-strand breaks occur at two specific sites associated with the hot spot and that occurrence of these breaks depends upon meiotic recombination functions RAD50 and SPO11. Furthermore, these breaks occur in a processed form in wild-type cells and in a discrete, unprocessed form in certain nonnull rad50 mutants, rad50S, which block meiotic prophase at an intermediate stage. The breaks observed in wild-type cells are similar to those identified independently at another recombination hot spot, ARG4. We show here that the breaks at ARG4 also occur in discrete form in rad50S mutants. Occurrence of breaks in rad50S mutants is also dependent upon SPO11 function. These observations provide additional evidence that double-strand breaks are a prominent feature of meiotic recombination in yeast. More importantly, these observations begin to define a pathway for the physical changes in DNA that lead to recombination and to define the roles of meiotic recombination functions in that pathway.