Single Cell Profiling of Cotton Ovules Epidermis During Early Fiber Development [scRNA-Seq]

Each cotton seed has approximately 25,000 singular fiber cells, which account for 25-30% epidermal cells. Understanding the basis of fiber cell initiation is critical for cotton yield improvement. However, it is difficult to separate and study fiber and non-fiber cells. Here we developed a reliable single-cell protocol to study transcriptome changes in Upland and Pima cotton and in a naked seed mutant. We analyzed over 40,000 single cells derived predominately from the epidermal layer of ovules during early stages of fiber development. Whole ovules of Gossypium were collected and briefly digested using macerozyme and cellulase enzyme mix to release protoplasts from the epidermal surrface, including developing fiber cells. Ovules from Gossypium hirsutum TM1 at 0, 1 and 2 days post-anthesis, the N1N1 fiberless mutant at 0 and 2 DPA and Gossypium barbadense at 2 DPA processed this way and used for 10x single-cell RNAseq.