Optimization of Cell Spreading and Image Quality for the Study of Chromosomes in Plant Tissues

• Published in: Methods in molecular biology (Clifton, N.J.) Vol. 1669; p. 141
• Main Authors: Kantama, Laksana, Wijnker, Erik, de Jong, Hans
• Format: Journal Article
• Language: English
• Published: United States 2017
• Subjects: Chromosomes, Plant - genetics; Euchromatin - genetics; Heterochromatin - genetics; In Situ Hybridization; Karyotyping; Cell spreading protocol; Image processing microphotographs; Chromosome technique; Chromosome images
• ISSN: 1940-6029
• DOI: 10.1007/978-1-4939-7286-9_12

High-quality chromosome images of mitotic and meiotic cell divisions in plant tissues are inextricably connected with the technical control of cell spread preparations. Superb chromosome slides are the best for studying chromosome morphology and making karyotypes; they also are the best start for a successful fluorescent in situ hybridization experiment. In this study, we describe the essentials for fixation, enzymatic digestion, squash, spread, and dropping protocols for clean and well-differentiated nuclei and chromosome complements. In addition, we focus on the use of standard whole image processing for best sharpness, brightness and contrast adjustments, differentiation of heterochromatin/euchromatin, and high dynamic range imaging of big chromosomes. We also explain how to combine transparent layers or spot channels of different fluorescent images for making publication quality, full color photos.