The dynamic behavior of semidilute polymer solutions is governed by an interplay between solvent quality, concentration, molecular weight, and flow type. Semidilute solutions are characterized by large fluctuations in polymer concentration, wherein polymer coils interpenetrate but may not be topologically entangled at equilibrium. In nonequilibrium flows, it is generally thought that polymer chains can “selfentangle” in semidilute solutions, thereby leading to entanglements in solutions that are nominally unentangled at equilibrium. Despite recent progress in the field, we still lack a complete molecular-level understanding of the dynamics of polymer chains in semidilute solutions. In this work, we use single molecule techniques to investigate the dynamics of dilute and semidilute solutions of k-phage deoxyribonucleic acid in planar extensional flow, including polymer relaxation from high stretch, transient stretching dynamics in step-strain experiments, and steady-state stretching in flow. Our results are consistent with a power-law scaling of the longest polymer relaxation time s ! ðc=c#Þ0:48 in semidilute solutions, where c is the polymer concentration and c* is the overlap concentration. Based on these results, an effective excluded volume exponent ! %0.56 was found, which is in good agreement with recent bulk rheological experiments. We further studied the nonequilibrium stretching dynamics of semidilute polymer solutions, including transient (1 c*) and steady-state (0.2 c* and 1 c*) stretching dynamics in planar extensional flow using an automated microfluidic trap. Our results show that polymer stretching dynamics in semidilute solutions is a strong function of concentration. In particular, a decrease in transient polymer stretch in semidilute solutions at moderate Weissenberg number (Wi) compared to dilute solutions is observed. Moreover, our experiments reveal a milder coil-to-stretch transition for semidilute polymer solutions at 0.2 c* and 1c* compared to dilute solutions. Interestingly, a unique set of molecular conformations during the transient stretching process for single polymers in semidilute solutions is observed, which suggests transient stretching pathways for polymer chains in semidilute solutions are qualitatively different compared to dilute solutions due to intermolecular interactions. Taken together, this work provides a molecular framework for understanding the nonequilibrium stretching dynamics of semidilute solutions in strong flows.