REFINED EXAMINATION OF PLANT METAPHASE CHROMOSOME STRUCTURE AT DIFFERENT LEVELS MADE FEASIBLE BY NEW ISOLATION METHODS

Two methods for isolation of plant metaphase chromosomes are described. The first, micromanipulation, allows the isolation of a number of individual chromosomes, which may be used as templates for the generation of chromosome specific DNA libraries and for physical sequence mapping by the polymerase chain reaction (PCR). The second provides, from synchronized meristems, pure chromosome suspensions suitable for flow cytometric analysis and chromosome sorting. Restriction endonuclease banding, immunostaining of chromosomal antigens, as well as fluorescence in situ hybridization at high signal to noise ratio were successfully performed on the isolated chromosomes. Chromosomes obtained by both protocols were suitable for scanning electron microscopy, the methods should also prove useful for refined analyses of the karyotypes of other plant species.