The role of rectification of heteroduplex heterozygotes in the formation of recombinant genotypes involving closely linked markers has been examined. Heteroduplex molecules of bacteriophage lambda DNA, heterozygous at several alleles, have been constructed and the genetic composition of phage present in infective centers derived by transfection with such molecules has been determined. Allele loss and concomitant recombinant formation is frequent, and appears to reflect marker specificity as well as specificities imposed by whether or not the transfection recipient is permissive or nonpermissive for DNA duplication of the transfecting genome. The observations support the proposal that many, perhaps most, of the events involving separation of closely linked markers occur by rectification of non-recombinant heterozygotes.