Low temperature and flooding are severe stresses to cotton plant growth and development in some parts of the world. This study was conducted to determine the interactive effects of low temperature and flooding on physiological and growth parameters of cotton plants, and yield of seed cotton. Transgenic Bt cotton plants at the boll-setting stage were treated with low temperature, flooding, and their combinations. The stresses decreased photosynthetic rate, stomatal conductance, transpiration rate, and intercellular CO₂ concentration by up to 37%, 71%, 52%, and 60%, respectively. The content of soluble protein and insecticidal protein was also decreased by up to 65% and 53%, respectively. By contrast, malondialdehyde (MDA) content increased by up to 114%. The length of fruiting branch was shortened by combined low temperature and flooding stress. Boll number and weight were decreased by 3.2% and 28.1%, respectively. Seed cotton yield was decreased by up to 43.2%. Our results suggest that concurrent low temperature and flooding at the boll-setting stage negatively influenced leaf photosynthetic activities and insecticidal protein production, leading to poor protection and yield. We recommend that agronomic measures be designed to stabilise physiological activities and insecticidal protein production in Bt cotton plants under stress conditions.