CHROMOSOMAL BAR CODES PRODUCED BY MULTICOLOR FLUORESCENCE INSITU HYBRIDIZATION WITH MULTIPLE YAC CLONES AND WHOLE CHROMOSOME PAINTING PROBES

Colored chromosome staining patterns, termed chromosomal 'bar codes' ( CBCs), were obtained on human chromosomes by fluorescence in situ hybr idization (FISH) with pools of Alu-PCR products from YAC clones contai ning human DNA inserts ranging from 100 kbp to 1 Mbp. In contrast to c onventional G- or R-bands, the chromosomal position, extent, individua l color and relative signal intensity of each 'bar' could be modified depending on probe selection and labeling procedures. Alu-PCR amplific ation products were generated from 31 YAC clones which mapped to 37 di fferent chromosome bands. For multiple color FISH, Alu-PCR amplificati on products from various clones were either biotinylated or labeled wi th digoxigenin. Probes from up to twenty YAC clones were used simultan eously to produce CBCs on selected human chromosomes. Evaluation using a cooled CCD camera and digital image analysis confirmed the high rep roducibility of the bars from one metaphase spread to another. Combina torial FISH with mixtures of whole chromosome paint probes was applied to paint seven chromosomes simultaneously in different colors along w ith a set of YAC clones which map to these chromosomes. We discuss the potential to construct analytical chromosomal bar codes adapted to pa rticular needs of cytogenetic investigations and automated image analy sis.