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Simple Summary: Coilia nasus is a migratory fish species with high nutritional and economic value, and naturally inhabits the middle and lower reaches of the Yangtze River and offshore China. Since the 1970s, the continuous deterioration of the habitat environment and continuous high-intensity fishing have led to a sharp decline in the germplasm resources of the Coilia nasus, which has been listed in the Red List of Threatened Species by the International Union for Conservation of Nature. Artificial farming has gradually become an important way to protect and utilize the germplasm resources of Coilia nasus, but limited germplasm sources and unscientific breeding strategies have put the germplasm resources and genetic diversity of Coilia nasus at risk, which limits its resources protection and utilization. The aim of this study was to assess the germplasm resources of Coilia nasus by analyzing the genetic diversity and genetic structure of its natural and farmed populations. In conclusion, the present study provides a reference for germplasm conservation and breeding strategy optimization in Coilia nasus, and contributes to the healthy development of Coilia nasus aquaculture. Coilia nasus is a threatened migratory species in the Yangtze River Basin. To reveal the genetic diversity of natural and farmed populations of C. nasus and the status of germplasm resources in the Yangtze River, the genetic diversity and structure of two wild populations (Yezhi Lake: YZ; Poyang Lake: PY) and two farmed populations (Zhenjiang: ZJ; Wuhan: WH) of C. nasus were analyzed using 44,718 SNPs obtained via 2b-RAD sequencing. The results indicate that both the wild and farmed populations had low genetic diversity, and germplasm resources have undergone varying degrees of degradation. Population genetic structure analyses indicated that the four populations may have come from two ancestral groups. Different amounts of gene flow were identified among WH, ZJ, and PY populations, but gene flow among YZ and other populations was low. It is speculated that the river–lake isolation of Yezhi Lake is the main cause of this phenomenon. In conclusion, this study revealed that genetic diversity reduction and germplasm resource degradation had occurred in both wild and farmed C. nasus, suggesting that conservation of its resources is of great urgency. This study provides a theoretical basis for the conservation and rational exploitation of germplasm resources for C. nasus. [ABSTRACT FROM AUTHOR]