The structures of λ DNA labeled during short pulses of [^3H]thymine at various times after infection have been studied by ultracentrifugal analysis. Circular DNA's, both the supercoiled form and an open form, are the main DNA components synthesized in the first half of the latent period. The open circular form is the most rapidly synthesized and may be a precursor to the supercoiled DNA. During the later period of progeny DNA accumulation the open circular form and a heterogeneous, more rapidly sedimenting component(s) are synthesized during such pulses. These are apparently involved in the generation of the linear, viral DNA