A METHOD FOR THE RAPID GENERATION OF ALPHA-SATELLITE AND CLASSICAL SATELLITE PROBES FOR HUMAN-CHROMOSOME-9 BY POLYMERASE CHAIN-REACTION USING GENOMIC DNA AND THEIR APPLICATION TO DETECT CHROMOSOMAL ALTERATIONSIN INTERPHASE CELLS

Fluorescence in situ hybridization (FISH) using chromosome-specific DN A probes is a technique which has recently become widely used for the analysis of chromosome alterations in interphase and metaphase cells, In this report, a polymerase chain reaction (PCR)-based method is desc ribed for simultaneously amplifying and labelling probes targeting the alpha- and classical satellite regions of chromosome 9 using either p lasmid or genomic DNA. Chromosome-specific probes were generated using readily obtainable plasmid DNA and genomic DNA from a hybrid cell lin e containing human chromosome 9 in a hamster cell background. The util ity of these probes to detect and quantify structural and numerical ab errations in interphase cells was demonstrated using a new multicolor FISH strategy by comparing the frequencies of hyperdiploidy and chromo some breakage affecting the regions targeted by the probes in interpha se and metaphase human lymphocytes irradiated during culture. The irra diated cells exhibited a significantly higher frequency of tetrasomy a nd breakage effecting the centromeric/pericentric region of chromosome 9 as compared with non-exposed cells, In general, similar frequencies of breakage and hyperdiploidy were observed in the interphase and met aphase preparations. These results show that DNA probes for the repeti tive sequences in human chromosomes can be easily generated from genom ic DNA and that these probes can be effectively used to detect chromos ome breakage and aneuploidy in interphase and metaphase lymphocytes in vitro.