Detection and calibration of microdeletions and microduplications by array-based comparative genomic hybridization and its applicability to clinical genetic testing

Abstract

Purpose: Genome-wide telomere screening by fluorescence in situ hybridization (FISH) has revealed that Ϸ 6% of unexplained mental retardation is due to submicroscopic telomere imbalances. However, the use of FISH for telomere screening is labor intensive and time consuming, given that 41 telomeres are interrogated. We have evaluated the use of array-based Comparative Genomic Hybridization (aCGH) as a more efficient tool for identifying telomere rearrangements. Methods: In this study, 102 individuals with unexplained mental retardation, with either normal or abnormal FISH results, were selected for a blinded retrospective study using aCGH. Results between the two methodologies were compared to ascertain the ability of aCGH to be used in a clinical diagnostics setting. Results:We detected 100% of all imbalances previously identified by FISH (n ϭ 17) and identified two additional abnormalities, a 10q telomere duplication and an interstitial duplication of 22q11. Interphase FISH analysis verified all abnormal array results. We also demonstrated that aCGH can accurately calibrate the size of telomere imbalances by using an array with "molecular rulers" for the telomeric regions of 1p, 16p, 17p, and 22q.Conclusion: This study demonstrates that aCGH is an equivalent methodology to telomere FISH for detecting submicroscopic deletions. In addition, small duplications that are not easily visible by FISH can be accurately detected using aCGH. Because aCGH allows simultaneous interrogation of hundreds to thousands of DNA probes and is more amenable to automation, it offers an efficient and high-throughput alternative for detecting and calibrating unbalanced rearrangements, both of the telomere region, as well as other genomic locations. Genet Med 2005:7(4):264 -271.

The evolution of molecular ruler analysis for characterizing telomere imbalances: from fluoresence in situ hybridization to array comparative genomic hybridization

“…11 These clones were used in pilot FISH studies to interrogate imbalances involving 1p, 6q, 10q, 16p, 17p, and 22q. 8,[11][12][13] Although these pilot studies only included a small number of cases per telomere, they demonstrated the utility of this high-density mapping for more accurate clinical interpretation of telomere imbalances. By determining the size of each telomere imbalance, pathogenic imbalances could be differentiated from those that were apparently benign variants, such as small deletions and duplications of 4q and 10q.…”

“…By determining the size of each telomere imbalance, pathogenic imbalances could be differentiated from those that were apparently benign variants, such as small deletions and duplications of 4q and 10q. 8,11 In this study, we have extended the use of FISH with our molecular ruler clone set to characterize a larger set of unbalanced telomere rearrangements. However, these analyses, as well as those of other investigators, 14,15 quickly led to the realization that many submicroscopic telomere imbalances were larger than 5 Mb and that additional bioinformatics and clone validation work to identify sequential clones for FISH mapping was inefficient and time-consuming.…”

Diagnostic Genome Profiling

“…In general, these arrays contain clones targeting genomic regions of known clinical significance, for example, all known microdeletion syndrome regions and all subtelomeric regions, which are known to be frequently affected in patients with mental retardation. [21][22][23] As a consequence, these arrays are very well suited for screening these specific regions but only these regions. Still, this is an important clinical application with considerable diagnostic yield in patients submitted for a variety of developmental problems, as was demonstrated by the recent publication of Shaffer et al 24 The diagnostic yield of targeted as well as genomewide microarray analysis, however, strongly depends on the clinical selection as well as the previously performed cytogenetic analyses, and this may vary considerably between countries and even between institutions within countries.…”

Fluorescence in situ hybridisation

“…The FISH studies using specific telomeric probes/clones have suggested that these probes are suitable diagnostic tool at examination of the mental retardation, infertility, congenital anomalies, at investigation of rearrangements in the haematological malign tumours, and in the pre-implanting diagnoses as well as in other fields of clinical interest Bacino et al 2000;Knight & Flint 2000;Caliskan et al 2005). The submicroscopic telomeric deletions, that are not well visible after FISH, could be, according to Wong et al (2005), detected more precisely by means of an array-based comparative genomic hybridization (CGH).…”