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This dissertation presents a simulation framework to investigate the capacity benefits and safety analysis for employing a proposed dynamic wake separation policy in a single lane flow corridor. The flow corridor concept is proposed as a Next Generation Transportation System (NextGen) route structure in en-route airspace to increase capacity in response to growing demand of air travel. To increase throughput, aircraft in flow corridors may fly closer to each other and have lower in-trail separations. But such aircraft must be safely separated with respect to wake vortices. Wake vortices are circular patterns of rotating air left behind a wing as it generates lift and can impose a significant hazard to other aircraft. Vortex trails, depending on atmospheric conditions, can persist for several minutes and many miles behind the generating aircraft in cruise altitudes. In this research, we consider a dynamic wake separation concept that uses information about actual weight and airspeed of aircraft and meteorological conditions to determine the minimum required wake separation between aircraft in a flow corridor. Aircraft characteristics and weather data are inputs to a fast-time wake prediction model that calculates the separation distances. These distances are updated periodically. To generate aircraft trajectories that are similar to real trajectories of aircraft in cruise, historical ADS-B flight track data are collected and analyzed. Trailing pairs in cruise altitudes are identified, and distributions for average and standard deviation of separation distance and ground speed, and standard deviation of altitude in level flight are obtained. Using these distributions, a simulation framework is developed to generate the trajectories of aircraft in a flow corridor. The simulation results demonstrate capacity benefits compared to current static separation standards. To demonstrate the safety of flow corridor operations, a rare event splitting methodology is used to estimate the probability of a potential wake encounter for a pair of trailing aircraft in cruise altitudes. Results of this simulation show that for aircraft trailing each other at the same altitude, occurrence of a potential wake encounter is very rare. Sensitivity analysis shows that altitude conformance is the most important parameter in determining the probability of the potential wake encounters. This analysis is extended to the flow corridor where every two consecutive aircraft can be considered as a trailing pair. Safety analysis is performed considering the worst-case scenarios that could happen in determining the dynamic separation, which demonstrate the safety of flow corridor in terms of wake vortex hazard.