To study the function of chalcone isomerase （CHI） gene in fiber development and response to stress in colored cotton，the physicochemical properties，conserved domain，cis-acting elements in the promoter of CHI gene，advanced structures and phylogeny of proteins were analyzed，transcriptome data and quantitative real-time PCR （qRT-PCR） were used to analyze its expression profile.The results showed that 12 members of GhCHI gene family were identified from the genome of upland cotton，which can be divided into two subfamilies with typical chalcone superfamily domains，encoding 201-452 amino acids，mainly hydrophilic proteins，and secondary structures mainly composed of α-helix and irregular curls.The types of cis-acting elements in the promoter of GhCHI gene mainly include light response elements，hormone response elements，and elements involved in the regulation of flavonoid biosynthesis.The results of analyzing expression profile showed that GhCHI1，GhCHI2，GhCHI3，and GhCHI4 were closely related to fiber development，especially with high level of expression during the period of pigmentation and coloring at the later stage of fiber development in colored cotton.These four genes also had high level of expression in leaves，receptacles，and pistils.GhCHI1，GhCHI2，and GhCHI3 were significantly induced to express under heat stress，salt stress，and drought stress.The results of qRT-PCR showed that GhCHI1-GhCHI4 were significantly enriched in fiber growth，especially in colored cotton.The results of analyzing the interaction of GhCHI1-GhCHI4 protein showed that they may interact with proteins such as 2-oxoglutarate 3-dioxygenase and flavonoid 3'- monooxygenase involved in the flavonoid synthesis.It is indicated that GhCHI1，GhCHI2，GhCHI3，and GhCHI4 genes play important roles in fiber development and stress response in colored cotton.