Sensitive and Rapid Detection of Centromeric Alphoid DNA in Human Metaphase Chromosomes by PNA Fluorescence In Situ Hybridization and Its Application to Biological Radiation Dosimetry

摘要

Centromeres of human chromosomes contain highly repeated sequences of DNA including alphoid DNA. Because of the complicated genomic organization of the centromere, the distribution of alphoid DNA in chromosomes has not been fully investigated. We conducted fluorescence in situ hybridization using a synthetic peptide nucleic acid as a sensitive probe (PNA–FISH) to detect chromosomal sites of alphoid DNA. As a result, the size variation of centromeric alphoid DNA among chromosomes was visualized with hybridization times as short as 1–2 h. In addition to the inter-chromosomal variation, we detected possible inter-individual variation in the size of alphoid DNA sites, which had been difficult to precisely analyze by conventional molecular and cytogenetic methods. We then applied this sensitive and rapid detection method to evaluate the yield of multicentric chromosomes induced in cultured human peripheral blood lymphocytes by high-dose gamma-irradiation. This PNA–FISH allows us to unequivocally determine centromeres in complexly rearranged chromosomes, confirming its usefulness in biological radiation dosimetry.