Schematic of the duplication region showing the relative positions of genes and FISH probes used (not to scale).

Molecular and cytogenetic data for patient 99.30. A. Genotyping with STS markers reveals evidence for an interchromosomal exchange leading to the formation of the der(15). Several STS in the proximal region between BP2-BP3 show only a single maternal allele, while distal to GABRB3 a biallelic maternal contribution is apparent. This region coincides with the area of increased dosage shown by array CGH (B). The array CGH data show tetrasomy of the region from BP1 that extends midway between BP2 and BP3. The distal end of this region through BP4 shows an increase in dosage to the hexasomy range. The interval between BP4 and BP5 is consistent with terasomy. The BP positions are indicated and the box indicates control probes. The positions of the clones used for FISH are highlighted. C-G. Metaphase FISH showing the der(15) chromosome indicated multiple signals for probes within the hexasomic region. Green signals are the centromeric probe pcm15. The BAC clones are indicated in red. H. Confirmation of hexasomy for clone AC127522 by interphase FISH with pcm15 (green) clone AC127522 (red). I. Schematic of the duplication chromosome for patient 99.30. The der(15) appears to have small satellites on each end. The der(15) extends to BP5 on each end but includes an two copies of an internally duplicated segment involving the distal portion of the BP2-BP3 interval and all of the BP3-BP4 interval indicated by the hatched yellow and green segments on the diagram. The position and orientation of the internally duplicated segment are not clearly defined but appear to be asymmetrically located within the der(15) based on the FISH data. In combination with two normal chromosome 15 homologs, this leads to hexasomy for the involved segments.

Molecular and cytogenetic analyses for 99.12, the mother of patient 99.10. A. Genotyping of the proband and parents identify a number of STS markers proximal to BP4 that are triallelic for the mother and tetra-allelic for the proband. The additional alleles suggest that the r(15) extends at least through BP4 and is present in the mother and proband. B. Log₂T/R ratios for the array CGH for the proband demonstrate tetrasomy for the region between BP1-BP4 and trisomy for the region between BP4 and BP5. This is consistent with a der(15) that arose from a BP4:BP5 exchange. C. Array CGH using peripheral white blood cells from the mother indicates a slight increase in dosage for clones between BP1 and BP5 (spots 1–73). The mean log₂T/R ratio was 0.108 (s.d. 0.060) compared with a mean ratio of 0.072 (s.d 0.065) (p = 0.037; two tailed unpaired t-test). For both arrays, the boxed area indicates control probes. The positions of the BAC clones used for FISH are indicated. D. Metaphase chromosomes hybridized with pcm15 (green) and BAC AC120045 (red) identifies two normal homologues and a small der(15) chromosome. E-J. Metaphase spreads hybridized with pcm15 (green) and additional BAC clones spanning the duplication regions. Most cells showed a single large signal for the pcm15 clone, suggesting the r(15) was monocentric.

Molecular and cytogenetic data for patient 00.16. A. Genotyping analyses using STS markers spanning the duplication region reveal evidence for an interchromosomal exchange leading to the formation of the der(15). The shift from biallelic to monoallelic maternal contribution could arise from a second exchange within the duplication or from a crossover on the normal maternal homologue. B. Graph of the Log₂T/R ratios for probes across chromosome 15q11-q14. The positions of the BP are shown as vertical lines. The dotted box indicates control probes distal to BP5 and from other autosomes. The positions of the FISH probes are indicated. The dosage data are consistent with tetrasomy of the region between BP2-BP3. C, Metaphase chromosomes hybridized with centromere probe, pcm15, and 770c6 show signals on the normal homologues as well as two centromeric signals on both der(15)s (inset). D and E. Images of idic(15) chromosomes from a single metaphase spread hybridized with pDJ437h9. F and G. Images of idic(15) chromosomes from a single metaphase spread hybridized with pDJ69i9. H. Schematic of the duplication chromosomes for subject 00.16. The der(15) der(15)s are satellited on both p arms, the heteromorphic region encompassing the NF1 pseudogene region is shown in light blue. The region between BP1-BP2 is shown in blue and present on both ends of the chromosome with a single copy of the region between BP2-BP3 shown in yellow. The approximate position of a potential crossover within the der(15) is shown by the dashed line.

Molecular and cytogenetic data for patient 03-46. A. Genotyping for the family of patient 03.46 shows biallelic maternal contribution for alleles proximal to BP3. Informative loci beyond BP3 have only a single maternal allele (D15S1019, D15S1043, D15S184). B. Array CGH shows an overall dosage from BP1-BP3 in the tetrasomic range. The dotted box indicates control probes and positions of the fish clones are indicated. C. Metaphase FISH using probes 142b22 (red) and AC026087 (green) identify the two normal homologues as well as a large der(15) with large confluent signals for the 142b22 and AC026087 probes (inset). D and E. Interphase FISH with the centromeric probe, pcm15 (blue), AC126407 (red) and AC026087 (green) identifies two cell lines, one euploid cell line (D) and one that carries a der(15) chromosome (yellow arrow) in addition to the two normal homologues (white arrows). The der(15) is dicentric and includes four signals for each of the BAC clones. F. Hybridization of interphase nuclei for 03.46 with pcm15 (blue), 142b22 (red) and AC024087 identifies two normal homologues (white arrows), one of which shows an expansion of the NF1/IgH pseudogene region with four signals from the 142b22 probe. The der(15) shows two centromeric signals, and eight signals from the 142b22 probe and four signals from the AC026087 BAC clone. Ordering of the signals suggests a head to tail arrangement for the duplicated segments within the der(15). G. Tissue section from 03-46 (case SS-99-5552) frontal cortex analyses by FISH for CEP15 (green) and GABRB3 (red). Nuclei were counterstained with DAPI (blue). Mosaicism of der(15) was observed in brain nuclei, with 60% of nuclei showing >2 spots per nucleus for CEP 15 or GABRB3 and 40% of nuclei showing ≤2 spots per nucleus for CEP15 and GABRB3. Homologous pairing of 15q11-q13 alleles was observed around the nucleolus in most nuclei, making the counts of FISH signals range from 1–4 spots per nucleus for CEP15 and 1–6 spots per nucleus for GABRB3 out of a total of 149 nuclei counted.