Protocol for the quantitative imaging and segmentation of spatial patterns in Hibiscus trionum petals

Most developmental processes underpinning the diversity of floral forms remain unexplored due to limitations in current imaging approaches. Here, we present a protocol to spatially analyze tissue-wide cell growth dynamics through confocal imaging and segmentation of the adaxial petal epidermis in Hibiscus trionum. We describe steps for dissecting buds, staining petals with a membrane dye, and acquiring images suitable for epidermal reconstruction. We detail procedures for image stitching and processing in ImageJ and for surface extraction and cell segmentation in MorphoGraphX. For complete details on the use and execution of this protocol, please refer to Riglet et al.¹.