Effective Chromosomal Preparation Protocol for the Dioecious Plant Silene latifolia

Cytology requires chromosome specimens, thus, effective preparation methods are needed. Chromosome specimens are frequently prepared from plant root tips. Furthermore, cell cycle synchronization using chemical reagents is applied to obtain a large number of metaphase chromosome specimens. In this study, we focused on the timing of root tip sampling, which is optimal for the preparation of chromosome specimens of the dioecious plant Silene latifolia. The timing was determined from the time seeds were subjected to a germination treatment. Observation of metaphase chromosomes using microscopy revealed that the number of mitotic cells peaked 54 h after the germination treatment. This trend was also observed when the DNA synthesis inhibitor aphidicolin was administered from 24 to 9 h before sampling time points. We used ice-cold treatment for 8, 16, and 32 h as a chromosome condensation method. The 16 h treatment produced suitable chromosome specimens showing satellite ends of chromosomes, whereas the 32 h treatment produced well-condensed chromosome specimens, which were suitable for counting chromosome numbers. Our findings suggest that the timing of root tip sampling is essential for effectively producing plant chromosome specimens.