Research on the evolutionary history of the morphological structure of cotton seeds: a new perspective based on high-resolution micro-CT technology

Cotton (Gossypium hirsutum L.) seed morphological structure has a significant impact on the germination, growth and quality formation. However, the wide variation of cotton seed morphology makes it difficult to achieve quantitative analysis using traditional phenotype acquisition methods. In recent years, the application of micro-CT technology has made it possible to analyze the three-dimensional morphological structure of seeds, and has shown technical advantages in accurate identification of seed phenotypes. In this study, we reconstructed the seed morphological structure based on micro-CT technology, deep neural network Unet-3D model, and threshold segmentation methods, extracted 11 basics phenotypes traits, and constructed three new phenotype traits of seed coat specific surface area, seed coat thickness ratio and seed density ratio, using 102 cotton germplasm resources with clear year characteristics. Our results show that there is a significant positive correlation (P< 0.001) between the cotton seed size and that of the seed kernel and seed coat volume, with correlation coefficients ranging from 0.51 to 0.92, while the cavity volume has a lower correlation with other phenotype indicators (r<0.37, P< 0.001). Comparison of changes in Chinese self-bred varieties showed that seed volume, seed surface area, seed coat volume, cavity volume and seed coat thickness increased by 11.39%, 10.10%, 18.67%, 115.76% and 7.95%, respectively, while seed kernel volume, seed kernel surface area and seed fullness decreased by 7.01%, 0.72% and 16.25%. Combining with the results of cluster analysis, during the hundred-year cultivation history of cotton in China, it showed that the specific surface area of seed structure decreased by 1.27%, the relative thickness of seed coat increased by 8.70%, and the compactness of seed structure increased by 50.17%. Furthermore, the new indicators developed based on micro-CT technology can fully consider the three-dimensional morphological structure and cross-sectional characteristics among the indicators and reflect technical advantages. In this study, we constructed a microscopic phenotype research system for cotton seeds, revealing the morphological changes of cotton seeds with the year in China and providing a theoretical basis for the quantitative analysis and evaluation of seed morphology.