We have used DNA-mediated gene transfer to study homologous recombination in the cells of higher eukaryotes. In the latest series of experiments we have introduced pairs of plasmid DNAs with mutations in the thymidine kinase gene of herpes simplex virus I (tk) into Ltk- mouse cells to demonstrate that the repair of double-strand gaps is a major mechanism for homologous recombination in these cells. We estimate that the L-cell can generate and repair a double-strand gap of between 152 and 248 base pairs. We concluded that double-strand gaps are recombinogenic in plasmid DNAs because they serve as intermediates in homologous recombination by double-strand gap repair, a gene conversion event. In another series of experiments we have been investigating the molecular mechanism of another recombination event: the human immunoglobulin gene class switch. We have obtained restriction mapping data for 6 switched human B cell lines indicating that the switching breakpoint in thr J-Cmu intron occurs within a small region at the 5' border of the mu "switch region" (Smu), these results are similar to those obtained by other investigators for switched mouse B cells. We have characterized and partially purified a recombinase from human cells. This activity catalyzes the formation of joint molecules from homologous double-strand linear (DS) and single strand circular (SS) DNA substrates. Up to 25% of substrate DNAs can be converted to joint molecules after 15 minutes of incubation at 37 degrees C. Joint molecules that have undergone strand exchange can be seen on electron micrographs of reaction mixtures. Homology between the DS and SS DNA substrates is required for the formation of joint molecules. Incubation of phix 174 SS DNA and M13 DS DNA (and vice versa) with extract does not result in formation of joint molecules.