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Gossypium hirsutum is an important source of natural textile fibers. Gossypol, which is a sesquiterpenoid compound mainly existing in the cotton pigment glands, can facilitate resistance to the stress from diseases and pests. The level of gossypol in the cotton is positively correlated to the quantity of pigment glands. However, the underlying regulatory mechanisms of gossypol synthesis and gland morphogenesis are still poorly understood, especially from a transcriptional perspective. The transcripts of young leaves and ovules at 30 DPA of the glanded plants and glandless plants were studied by RNA-Seq and 865 million clean reads were obtained. A total of 34,426 differentially expressed genes (DEGs) were identified through comparative transcriptome analysis. Genes related to gossypol synthesis or gland morphogenesis displayed significant differential expression between the two cultivars. Functional annotation revealed that the candidate genes related to catalytic activity, the biosynthesis of secondary metabolites, and biomolecular decomposition processes. Our work herein unveiled several potential candidate genes related to gossypol synthesis or gland morphogenesis and may provide useful clues for a breeding program of cotton cultivars with low cottonseed gossypol contents. [ABSTRACT FROM AUTHOR]