Abstract: Gymnocypris przewalskii, an endemic migratory fish species in Qinghai Lake and its surrounding waters, is capable of adapting to the lake’s highly saline-alkaline environment. The solute carrier (SLC) family, as key membrane transport proteins, participates in numerous essential physiological and metabolic pathways and may play critical roles in the adaptation of G. przewalskii to saline-alkaline stress. To systematically elucidate the function and regulatory mechanisms of SLC41A1 in G. przewalskii, we first retrieved its sequence from the G. przewalskii genome database and cloned the coding sequence (CDS) and promoter region. We analyzed the structural features of the encoded protein, its tissue-specific expression patterns, and its transcriptional response to saline-alkaline stress. Subsequently, bioinformatic analyses were employed to predict core promoter regions, regulatory elements, and transcription factor binding sites. Finally, a dual-luciferase reporter assay was used to assess the transcriptional activity of various truncated promoter fragments. The CDS of SLC41A1 gene in G. przewalskii is 1047 bp in length, encoding a protein of 348 amino acids that contains transmembrane transport-related domains and exhibits high conservation among teleost fishes. Under saline-alkaline stress, SLC41A1 expression was significantly upregulated in kidney and intestinal tissues, indicating its role in environmental adaptation. A 2148-bp promoter fragment of SLC41A1 was obtained, within which a core promoter region and three potential key transcription factor binding sites (C/EBPα1p, HNF-1, and Sp1) were identified. Transcription assays revealed strong transcriptional activity in the region from –334 to –555 bp upstream of the transcription start site, likely due to the dense clustering of binding sites for C/EBPα1p, HNF-1, and Sp1. This study provides new insights into the potential role of SLC41A1 gene and its regulatory sequences in saline-alkaline adaptation, offering a foundation for further understanding the molecular mechanisms underlying G. przewalskii’s adaptation to the unique high-altitude environment of the Qinghai-Xizang Plateau.