Effects of acetic acid treatment on plant chromosome structures analyzed by atomic force microscopy

Acetic acid treatment has been frequently used to remove cellular contaminants from plant chromosome samples for structural analyses by scanning electron microscopy and atomic force microscopy (AFM). We evaluated the effects of various concentrations of acetic acid treatments on barley chromosome structures by using AFM. The long-term 45% acetic acid treatment significantly damaged the chromosome structures, although the treatment effectively removed the cellular contaminants. On the other hand, the treatment with 15% acetic acid could not obtain sufficiently clean chromosome samples and the chromosome surface structures could not be observed. In contrast, we obtained clean chromosome preparation without severe damage by using an intermediate concentration (30%) of acetic acid treatment. In the centromeric region, we could observe fiber structures with a width of 100 nm, which were composed of ca. 50-nm granules and aligned to the axes of chromosomes. Thus, AFM analysis of chromosomes appropriately treated with acetic acid will provide important insights into the organization of higher-order structures of plant chromosomes.