Comparative evaluation of maize chromosome pretreatment using cold water and 8-hydroxyquinoline

In this study, a more efficient and time-saving method for chromosome preparation in maize is proposed. Studying chromosomes is crucial for understanding their organization, abnormalities (such as translocations, bridges, and aneuploidy), structure, and the evolution of maize genes and genomes. While there are several valuable protocols available for chromosome preparation-for purposes like counting chromosomes and performing karyotype analysis-these methods are often time-consuming, and the proportion of metaphase cells can be low. Therefore, it is important to explore more efficient methods for preparing maize chromosomes. Pretreatment plays a crucial role in chromosome preparation. This study compared the effects of different pretreatment methods (cold pretreatment and 8-hydroxyquinoline pretreatment), varying durations of pretreatment (cold pretreatment: 6, 12, 24, and 36 h; 8-hydroxyquinoline pretreatment: 0.5, 1, 2, 3, 4, and 5 h), and different fixation durations (1, 2, 3, 6, 12, 16, 20, 24, and 28 h) on maize chromosome preparations (Zea mays L.). The results indicated that cold pretreatment lasting over 12 h yielded the most countable chromosome figures. Additionally, pretreatment with 0.006 M 8-hydroxyquinoline for 2-3 h showed no significant difference in the number of countable chromosome figures. However, the proportion of mitotic cells significantly decreased when treated for four or five h with 0.006 M 8-hydroxyquinoline. Furthermore, there was no significant difference in the proportion of metaphase cells when fixed with Carnoy's fixative, regardless of the fixation time used. Based on these findings, a more efficient and time-saving method for chromosome preparation in maize is proposed: the root tips of maize should be pretreated with a mixture of ice and water at 4 °C for 12 h or with a 0.006 M 8-hydroxyquinoline solution at room temperature for 3 h. The root tips should then be fixed with Carnoy's fixative (composed of glacial acetic acid and absolute ethanol in a 1:3 ratio) for 1 h.