Identification of the origin of double minutes in normal human cells by laser-based chromosome microdissection...

Single copies of tiny chromosome fragments, appearing as double minutes, were observed in a high proportion of cells from amniotic fluid cultures of two mothers undergoing prenatal testing because of advanced age. We applied a laser-based chromosome microdissection method to diagnose the origin of the double minutes. The diagnostic procedures consisted of microdissection of double minutes from a single cell, polymerase chain reaction (PCR) amplification of the dissected DNA, and subsequent fluorescence in situ hybridization (FISH) using the PCR products as a probe pool. Metaphase chromosomes from the patients' cells and from a karyotypically normal individual were probed. Using this strategy, we were able to determine that the double minutes originated from the centromere of chromosome 13 or 21 in one case, and from the chromosome 12 centromere in the other. The characterization of such double minutes helps both in the delineation of the nature of these epichromosomal bodies in normal individuals as well as in the clarification of genetic counselling issues.