Application of Micro-FISH for Characterization of Structural Human Chromosome Abnormalities

Structural chromosome aberrations, especially de novo translocations and marker chromosomes, often remain uncharacterized in clinical cytogenetic analysis. Due to the limitations of conventional banding analysis in precisely identifying these rearrangements and marker chromosomes, genetic counselling is exceedingly difficult. Microdissection combined with fluorescence in situ hybridization (micro-FISH) has become a powerful tool in clinical genetics for the characterization of cytogenetically unclassifiable aberrations. Micro-FISH was used to elucidate the chromosomal origin of two different de novo structural chromosome abnormalities. Ten copies of aberrant chromosomes were collected with microneedles from patient's metaphases, transferred to a collecting drop and amplified by means of degenerate oligonucleotide primed polymerase chain reaction (DOP-PCR). The PCR products were labeled by nick-translation with digoxigenin-11-dUTP and used as FISH probes. They were hybridized to metaphase spreads from patients to confirm the specificity of the probe and normal metaphases to determine the origin of the aberrant chromosomes. With this strategy, a de novo marker chromosome and a de novo isodicentric chromosome in peripheral blood samples were successfully identified in two cases. One marker of a small ring chromosome appeared to be derived from the pericentromeric region on the short arm and long arm (Xp11.1-q12) of the X chromosome and the second aberrant was identified as an isodicentric X chromosome (idic(X)(q28)). Based on the analysis of both G-banding and micro-FISH, the karyotypes for the patients were defined as mos 46,X,r(X)(p11.1q12)(64)/45,X(26)/46,XX(10) for the first patient and mos 46,X,idic(X)(q28)(86)/ 45,X(12)/46,XX(2) for the second case, respectively.