SRA

Comprehensive utilization of cottonseeds is limited by the presence of pigment gland and its inclusion gossypol. The ideal cotton is glandless-seeds and glanded-plant, a trait found in only few Australian wild cotton species, including Gossypium bickii. Introgressing the trait to cultivated species is proved to be difficult. Understanding the biological processes towards pigment gland morphogenesis and the associated underlying molecular mechanisms will facilitate breeding cultivated cotton varieties with the trait of glandless-seeds and glanded-plant. Single-cell RNA sequencing (scRNA-seq) was performed on 12,222 protoplasts isolated from cotyledons of germinating G. bickii seeds 48-hours after imbibition. Clustered into 14 distinct clusters unsupervisedly, these cells could be grouped into eight cell populations with the assistance of known cell marker genes. The pigment gland cells were well separated from others, and could be separated into pigment gland parenchyma cells, secretory cells, and apoptotic cells. In this study, integrating pigment gland cells developmental trajectory, transcription factors regulatory networks, and core transcription factors functional validation, a relatively complete model was proposed for pigment gland formation. Light and gibberellin were verified to promote the formation of pigment glands. Besides, three novel genes, GbiERF114 (ETHYLENE RESPONSE FACTOR 114), GbiZAT11 (ZINC FINGER OF ARABIDOPSIS THALIANA 11) and GbiNTL9 (NAC TRANSCRIPTION FACTOR-LIKE 9), were found to affect pigment gland formation. These findings shed new insights into pigment gland morphogenesis and lay the cornerstone for future cotton scRNA-seq investigations. Overall design: Comparative gene expression profiling analysis of bulk RNA-seq data along the germination and under the treatments of dark and light of G. bickii cotyledons