Abstract: In recent years, high temperatures have become an important abiotic stress factor that limits the growth and development of cotton in the Yellow River basin. The characteristics of different cotton cultivars in response to high temperatures, especially differences in root phenotypes, remain unclear. In this study, 15 cotton cultivars commonly cultivated in the Yellow River basin were cultured to the six-leaf stage in an artificial climate chamber under normal conditions (25 ℃ day/25 ℃ night), followed by treatments with control (25 ℃ day/25 ℃ night) and high (35 ℃ day/30 ℃ night) temperatures. Seven days later, 10 physiological indicators, including gas exchange parameters, chlorophyll fluorescence parameters, antioxidant system enzymes, and root phenotypic parameters, such as root length, root surface area, root volume, and average root diameter, were measured. The results showed that, compared with the control, for all cultivars, values of net photosynthetic rate, stomatal conductance, transpiration rate, PSⅡ maximum photochemical efficiency (F_v/F_m), and relative chlorophyll content (SPAD) generally decreased after high-temperature treatment; while activities of superoxide dismutase (SOD), peroxidase (POD) and catalase (CAT), soluble sugar content, and relative conductivity increased. Ten indicators were integrated into two comprehensive indicators, and the high-temperature resistance coefficient and cultivar score for each cultivar were obtained using principal component analysis. Three high-temperature-resistant cultivars, ‘Shuofeng 1’ ‘Guoxin 9’ and ‘Lumianyan 28’; and five high-temperature-sensitive cultivars, ‘Shikang 126’ ‘Hanwu 216’ ‘Guoxin 4’ ‘Cangmian 268’ and ‘Nongda 601’, were screened out via cluster analysis. The correlation between the ratio of high temperature to control of root phenotypic indicators and the high-temperature tolerance score of high-temperature-resistant cultivars was further analyzed. The correlation coefficients of root length, root surface area, root volume, and mean root diameter were 0.766 (P<0.01), 0.659 (P<0.01), 0.628 (P<0.05), and 0.501 (P>0.05), respectively, indicating that root phenotypic parameters decreased less after high-temperature stress. Root length, surface area, and volume can also be used as indicators to screen cultivars resistant to high temperatures. This study provides theoretical and practical support for the selection and regulation of high-temperature-resistant cultivars.