Torsionally unconstrained double-stranded DNA undergoes an “overstretching” transition at around 65 pN which leads to DNA elongation of ~1.7-fold. Recent experiments have revealed that it involves two distinct structural transitions: a “strand-unpeeling” of one strand from the other, and a “B-to-S” transition to a mysterious double-stranded “S-DNA”. Here, we report the first simultaneous determination of the entropy and enthalpy changes, as well as whether the two DNA strands are spatially separated, during the respective transitions. Quantitative understanding of the selection of the two transitions is illustrated in phase diagrams including B-DNA, single-stranded DNA, and S-DNA. Our results provide important insights in the thermodynamics of DNA overstretching and the structures of overstretched DNA that might play an important role in vivo.