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The upper reaches of the Yangtze River (upper YR) are a biological zone with extremely rich fish diversity, especially endemic fish. However, long-term human interference, such as environmental pollution and cascade hydropower construction, has significantly changed the habitat of many fish and is threatening the number and genetic diversity of fish populations. <i>Jinshaia sinensis</i> is a typical small and endemic but rare fish that is found in the upper YR, and its genetic diversity and structure still need further study. To understand the current levels of genetic diversity in <i>J. sinensis</i>, we analyzed the genetic diversity, population history, genetic structure, etc., of three <i>J. sinensis</i> populations based on two mitochondrial genes (the cytochrome-c oxidase subunit I, COI, and cytochrome-b gene, Cytb) and two nuclear genes (recombination-activating protein 1, RAG1, and rhodopsin, RH). The genetic diversity analysis indicated that <i>J. sinensis</i> had high genetic diversity, with high haplotype diversity (<i>h</i>) and nucleotide diversity (Pi). Population pairwise F_ST analysis revealed a significant genetic divergence between the Lijiang and Luzhou populations for all genes and between the Panzhihua and Luzhou populations, except for the COI gene; however, analyses of molecular variance (AMOVA) showed no significant geographic genetic structure among populations, and gene flow analysis also indicated a certain degree of gene exchange among populations. Haplotype network structure analyses revealed low levels of shared haplotypes among populations. Neutrality test and mismatch distribution results indicated that only the Lijiang population had experienced obvious population expansion. Overall, these results indicate that <i>J. sinensis</i> is still a single evolutionarily significant unit, but when considering the threat of habitat disturbance to the population, it is still necessary to carry out long-term genetic monitoring on <i>J. sinensis</i> and on other endemic fishes with similar ecological habits in order to maintain the genetic diversity of fishes in the upper YR.