HYBRIDIZATION BASED KARYOTYPING: HYBRIDIZATION BANDS

Applied Genetics Laboratories, Inc., 1335 Gateway Drive, Suite 2001, Melbourne, FL 32901 USA

We report a method, Hybridization-banding, (H-banding) that allows simultaneous identification of all the chromosomes in metaphase spreads. The method uses a combination of hybridization probes labeled with a single fluor to yield a simple, unique, readily identifiable pattern of hybridization on every chromosome in one hybridization procedure. H-banding is superior to Giemsa or fluorescence based banding methods for chromosome identification in that the simpler hybridization patterns produced are easier for inexperienced personnel to identify. Because the method does not use multiple fluors or fluor combinations, analysis can be performed without specialized equipment. Instead, analysis can be performed with a standard fluorescence microscope and images recorded on film with an ordinary 35-mm camera, making the method useful to many investigators. The method can be used to readily identify and quantitate chromosomes. Further, the method can be used to quickly map any DNA probe to the proper chromosome by labeling that probe in a second color and including it in the hybridization. Our scheme for mouse combines 27 cloned point probes and 6 band-specific probes derived from microdissection to identify every chromosome, but other effective combinations can be devised. Unique patterns can be designed at will for each chromosome, allowing H-banding to be tailored for specific needs. Further, the method can also be applied to any animal or plant species for which chromosomes and probes can be prepared.