Fluorescent in situ hybridization of maize meiotic chromosomes as visualized by confocal microscopy

Knob heterochromatin served as the model for the development of fluorescent chromosome in situ hybridization on maize meiotic chromosomes. The meiotic chromosomes were hybridized with a digoxigenin-labeled RNA probe of the knob repeat sequence that is a component of the morphologically determined knob heterochromatin. The fluorescein-labeled knob probe and propidium iodide counter-stained chromosomes were imaged using confocal laser scanning microscopy, which allowed for the individual analysis of each fluorescent probe emission intensity, the ability to utilize image processing techniques and the generation of high-resolution images. A composite, in register, merged image of the knob probe signal and meiotic chromosomes demonstrated exact co-localization of the knob probe and the morphologically identified knobs. The establishment of the fluorescent in situ hybridization technique in maize allows for the expanded study of the biological role of knob heterochromatin and the possibility of locating other repeat sequences on maize chromosomes.