Abstract: Takifugu species (order: Tetradotiformes; family: Tetradontidae) have attracted attention among the scientific community for their peculiar biology, physiology and genomics. Obscure puffer (Takifugu obscurus) is an anadromous fish species in China. They usually grow in the sea before turning sexually mature, then migrate to freshwater rivers for reproduction during the spawning season. Obscure puffer is a popular fish in China for its high-quality meat. In 2002, annual production of obscure puffer was 2600 tons, and it has been continuing to rise owing to its large body size, rapid growth, and high market value of the fish. However, obscure puffer resource has sharply declined because of overfishing and water pollution. Previous studies have focused on the physiology and mitochondrial DNA. Analysis of the genetic diversity and structure of a fish species is an important prerequisite for conservation as it reflects the status and survival potential of this species. However, the genetic diversity among and within populations of obscure puffer populations remains unknown, thus it is difficult to implement effective conservation strategies. Molecular markers are useful for population genetic studies to assess the influence of various factors on genetic diversity and population structure. Among the many types of molecular markers, sequence-related amplified polymorphism (SRAP) has been recognized as a new and useful molecular marker system because of its simplicity, reproducibility and convenience. This method is based on two-primer amplification. The primers are 17 or 18 nucleotides long and consist of the following elements. SRAP amplifies open reading frames and is significant for gene and genetic diversity. It has been applied extensively in molecular identification, genetic linkage map construction, gene tagging, genetic diversity analysis and comparative genetics of different species.In this study, in order to investigate the genetic diversity and population structure among and within populations of obscure puffer, a SRAP marker was firstly used to analyze the genetic diversity of four obscure puffer populations. These populations included a captured population in the Yangtze Rive, a released population and two aquaculture populations. 18 pairs of primer combinations, which were selected from 49 primer pairs, were applied to four populations. A total of 231 loci were detected, among which 156 were polymorphic. The proportion of polymorphic loci ranged from 54.98% to 58.87% in these four populations. The Nei’s genetic diversity (H) of the four populations were between 0.1992 and 0.2005, while the Shannon information index(I) were between 0.2953 and 0.3016. The gene flow (Nm) was 4.1291 among populations, of which, the proportion of polymorphic loci, the Nei’s genetic diversity(H) and Shannon information index (I) of the captured population were a little higher than the other three populations. Individual clustering showed that the Shanghai population formed an independent cluster and the other three populations belonged to another cluster. AMOVA analysis showed that 87.40% of variance was within population. These genetic statistics indicated there were a richer genetic diversity in four populations and a certain gene flow between them.