Seagrasses are declining at an alarming rate in Florida, prompting increased restoration efforts throughout the state. Literature on other organisms highlights the importance of selecting restoration materials with natural resiliency and resistance to stressors, which may enhance overall success. Currently, many restoration managers, including those in the Florida Aquatic Preserves (AP), use Halodule wrightii as restoration material due to its natural abundance and importance to the marine meadow ecosystem. While some genetic research has been conducted using microsatellite markers on H. wrightii, there are no studies along Florida’s Gulf Coast employing 2b-RAD sequencing, which can provide information on genetic diversity and connectivity, as well as historical reconstructions of population divergence. For this study, we collected samples that were genotyped using microsatellite markers and combined them with novel samples from several locations along Florida’s Gulf Coast to determine the current genetic diversity, connectivity, and historical population divergences across geographic regions. This data has the potential to assist conservation efforts in Florida’s declining seagrass populations.