Cell-cycle synchronization and flow karyotyping in rice

Highly efficient cell synchronization and metaphase chromosome accumulation in rice root-tip cells were achieved. Flow cytometric analysis was performed for obtaining optimal parameters to synchronize the cell cycles. High mitotic indices (57.6% in root meristematic region) were obtained by treating 0.5-cm seedlings with 0.5 mM hydroxyurea at 30 oC for 4 h, incubating in a hydroxyurea-free solution for 30 min, and then treating with 0.3 µM trifluralin for 3 h. After trifluralin treatment, incubation in distilled water for 15 min reduced chromosome clumping on metaphase spread. Uniformity in seed germination at the time of treatment is a critical parameter for obtaining a high metaphase index. Isolated rice chromosomes were suitable for flow cytometric analysis and chromosome sorting. The morphology of flow-sorted metaphase chromosomes was not affected.