A new approach to analysis of complex chromosomal rearrangements in cell hybrids

The chromosomal complements of somatic cell pig-mink hybrids was determined by a new approach. This approach includes microdissection of metaphase chr omosomes, generation of chromosome and region-specific DNA libraries, and f luorescence in situ hybridization of these libraries with pig lymphocyte ch romosomes. The studied hybrid cells were shown to contain two small acrocen tric chromosomes and a microchromosome of porcine origin. Identification of these chromosomes by differential GTG-staining was impossible. Chromosome isolation by a micromanipulation technique followed by DNA amplification in TOPO-DOP polymerase chain reaction provided chromosome-specific DNA librar ies of the rearranged chromosomes. Based on these libraries, the labeled DN A probes were prepared and hybridized to pig chromosomes. This allowed us t o determine the origin of the material contributing to the hybrid cell chro mosomes. One of these chromosomes contained five pig chromosomal regions: 1 5cen-q2; 6q21-q23; 13q21; 13q22; 7q25-qter, while the other contained the f ollowing pig chromosomal regions: 4p12-p13; 16q12-q14; 12pter-p15. The micr ochromosome contained the Xp11-Xq11 region. The minimal size of the reveale d chromosomal regions was about 3 to 4 x 10(6) bp. Segregation analysis of the thymidine kinase gene 1 (TK1), which was earlier localized to the pig 1 2p region, and the hybrid cell pig chromosomes in the hybrid, subclones sug gested that TK 1 gene can be assigned to 12p15-pter. The results obtained d emonstrate the efficiency of the applied approach in its detailed and relia ble description of complex chromosomal rearrangements in hybrid clones, whe n differential chromosome staining failed to identify these chromosomes.