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Rat and mouse have become important animal models to study various human diseases such as cancer. Cytogenetic analysis of the respective karyotypes is frequently required to investigate the causative genetic defects and especially neoplastic cells often show complex chromosome aberrations and many different marker chromosomes. However, structural homogeneity of the chromosomes in these species as well as less pronounced differences in banding patterns make it difficult to assign genetic abnormalities to certain chromosomes by conventional banding techniques. Here we report for the first time the successful application of multicolor spectral karyotyping (SKY) to rat chromosomes, which allows unequivocal identification of all rat chromosomes with the exception of chromosomes 13 and 14 in different colors, thus enabling the elucidation of even complex rearrangements in the rat karyotype. Flow-sorted chromosome specific painting probes for all 22 rat chromosomes (20 autosomes, X, and Y) were combinatorially labeled by a set of five different fluorochromes and hybridized in situ to metaphase spreads of a healthy rat, to diakineses from testicular material, and to cells from a rat FAO hepatoma cell line. Measuring the complete spectrum at each image point by using the SpectraCube((R)) spectral imaging system and respective computer software allowed identification of the individual rat chromosomes by their specific emission spectra. Classification algorithms in the analysis software can then display the rat chromosomes in specific pseudo-colors and automatically order them in a karyotype table. After its successful application to human and mouse chromosomes, spectral karyotyping of rat chromosomes now also allows cytogenetic screening of the complete rat genome by a single hybridization., (Copyright 2003 S. Karger AG, Basel)

A technique disclosing most information about chromosome modifications is the technique of choice for the analysis of chromosome alterations. The newly developed method for microdissection of fluorescence-labeled chromosomes (FISH-MD) can improve upon this expectation in combination with 24-color spectral karyotyping (SKY). The highly efficient way to detect chromosome modifications by SKY and the detailed specification of aberrant chromosomes by FISH-MD prompted us to use both techniques in a combined approach called SKY-MD. First, an overview of chromosomal aberrations is obtained by spectral karyotyping and subsequently the derivative chromosomes recognized are characterized in a highly specific manner by microdissection and reverse painting. A small quantity of isolated material dissected directly from a 24-color metaphase is sufficient to obtain very detailed information about the chromosome regions and the breakpoints involved in the derivative chromosomes. Therefore, the combination of spectral karyotyping and microdissection in one procedure, and reverse painting can characterize chromosomal aberrations with a degree of specificity hitherto unknown from individual karyotyping experiments. In this article we compare the efficiency of both the SKY technique and that of classical microdissection with the efficiency obtained by SKY-MD.