Preparation of Mitotic Chromosomes with the Dropping Technique

Abstract

The quality of chromosome preparation influences all downstream analyses and is therefore crucial. Hence, numerous protocols exist to produce microscopic slides with mitotic chromosomes. Nevertheless, due to the high content of fibers in and around a plant cell, preparation of plant chromosomes is still far from trivial and needs to be fine-tuned for each species and tissue type. Here, we outline the "dropping method," a straightforward and efficient protocol to prepare multiple slides with uniform quality from a single chromosome preparation. In this method, nuclei are extracted and cleaned to produce a nuclei suspension. In a drop-by-drop manner, this suspension is then applied from a certain height onto the slides, causing the nuclei to rupture and the chromosomes to spread. Due to the physical forces that accompany the dropping and spreading process, this method is best suited for species with small- to medium-sized chromosomes.

Keywords: Chromosome preparation; Dropping technique; Karyotyping; Metaphase preparation; Microscopy; Molecular cytogenetics; Plant chromosomes.