Abstract: Taking the cold-tolerant Carya illinoinensis from Xinjiang Provenance (XJ) and cold-sensitive “Shaoxing No.1”C. illinoinensis (SX) half-sib families of C. illinoinensis as research subjects, this study compared the physiological and biochemical changes in leaves under low-temperature treatment, including superoxide dismutase (SOD) activity, proline (Pro) content, and malondialdehyde (MDA) content. Through transcriptome sequencing, the changes in gene expression in leaves before and after low-temperature treatment were analyzed. The findings indicated that after low-temperature treatment, compared with the control, the physiological indicators in response to low-temperature, SOD, proline, and MDA, increased by 58.11%, 39.30%, and 3.42% respectively in XJ, while in SX, they increased by 152.53%, 94.52%, and 7.97% respectively. Under low-temperature treatment, genes related to oligosaccharide biosynthesis and metabolism were upregulated in the XJ family of C. illinoinensis, whereas genes involved in low-temperature-related cell surface receptor and membrane receptor signal transduction pathways, as well as those associated with meristem and growth and development, were suppressed. Under low-temperature stress, genes involved in brassinosteroid biosynthesis and signal transduction pathways were significantly enriched among the upregulated genes in the XJ family compared with the SX family. Therefore, the XJ family of C. illinoinensis may possess a genetically enhanced capacity for oligosaccharide synthesis. Under low-temperature stress, XJ also participates in the low-temperature stress response by enhancing the biosynthesis and signaling of brassinosteroids and increasing gene expression in the CBF pathway, thereby improving its cold resistance.