Abstract: 【Objective】 Exploring the mechanism of cotton tolerance to salt and alkali stress plays an important role in improving cotton yield in Alkali soil in Xinjiang. 【Methods】 This study used cotton as the experimental material, and set up three treatments: control (CK), salt stress (NaCl, CS), and alkali stress (NaHCO₃+Na₂CO₃, AS). Then, TMT technology was used to analyze the changes in protein expression in cotton roots under salt and alkali stress, and bioinformatics analysis was performed on the screened differential proteins. 【Results】 The results showed that under salt stress, the activities of Hexokinase, Phosphofructokinase, Pyruvate kinase and citrate synthase in roots were significantly reduced by 12.0%, 9.9%, 9.7%, and 32.8%, respectively. Under alkali stress, the activities of Hexokinase, Phosphofructokinase, Pyruvate kinase, Malate dehydrogenase, Glutamate dehydrogenase and cereal Transaminase in roots were significantly increased by 10.9%, 5.9%, 10.4%, 45.1%, 26.3%, and 23.4%, respectively. A total of 1725 differentially expressed proteins were screened under salt stress, including 508 upregulated differentially expressed proteins and 1217 downregulated differentially expressed proteins. Among them, there were 10 upregulated differentially expressed proteins and 43 downregulated differentially expressed proteins in energy metabolism, 14 upregulated differentially expressed proteins and 76 downregulated differentially expressed proteins in amino acid metabolism, 45 upregulated differentially expressed proteins and 84 downregulated differentially expressed proteins in genetic information processing, and 13 upregulated differentially expressed proteins and 29 downregulated differentially expressed proteins in signal transduction; Under alkaline stress, there were a total of 75 differentially expressed proteins, including 30 upregulated differentially expressed proteins and 45 downregulated differentially expressed proteins. Among them, there were only 2 downregulated differentially expressed proteins in energy metabolism, 1 upregulated differentially expressed protein in amino acid metabolism, and 3 downregulated differentially expressed proteins. No differentially expressed proteins were identified in genetic information processing and signal transduction. 【Conclusion】 The results indicate that salt stress significantly inhibits energy metabolism, amino acid metabolism, genetic information processing and signal transduction in cotton roots, while alkali stress has no significant effect on genetic information processing and signal transduction.