<br> The ecological opportunities presented by the invasion from marine to freshwater habitats are considered to be an important catalyst for the adaptive radiation of many fish taxa. The sculpins (family Cottidae) serve as a typical representative of such successful evolutionary radiation from marine to freshwater environments, and among which, several intermediate diadromous taxa that can adapt to both marine and freshwater habitats have been evolved. However, little is known about the genetic basis underlying the diadromous migration adaptations for these euryhaline sculpins. Here, we have constructed a high-quality chromosome-level genome of the Cottidae species roughskin sculpin (<br> Trachidermus fasciatus<br> ) and made a comparative genomic analysis of the genomes from the closely related species, aiming to elucidate the genetic basis and the possible evolutionary origin of its unique diadromous lifestyle. Our Hi-C data anchored the preliminary assembled sequences to 20 pseudochromosomes, resulting in a final corrected genome size of 526.39 Mb, with a scaffold N50 of 24.94 Mb. Phylogenetic analysis supports that the Cottidae family, to which roughskin sculpin belongs, possibly originated from the ocean, and should be reclassified under the suborder Cottoidei of the order Perciformes, rather than the classical order Scorpaeniformes. Comparative genomic analysis showed that 184 and 856 putative gene families in roughskin sculpin have undergone expansion and contraction, respectively. Enrichment categories of the top candidate genes under significant expansion were mainly associated with oxygen transport and immune response, possibly indicating their adaptations to the great oxygen demand and a wider range of pathogens across different salinity gradients during diadromous migration of roughskin sculpin. Additionally, 477 putative genes were identified to have undergone positive selection, which were enriched in categories related to DNA repair, innate immunity, and ion transport. This may indicate that the adaptive remodeling of immune systems and osmotic regulation mechanisms is essential for the roughskin sculpin to cope with the challenges posed by exposure to diverse microbial communities and fluctuations in salinity during migration. Our work not only provides valuable resources for understanding the genetic basis of the roughskin sculpin’s adaptation to a unique migratory lifestyle, but may also offer important insights into how evolutionary radiation from marine to freshwater habitats has been achieved in marine-originated sculpins.<br>
