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Preimplantation Genetic Testing for Aneuploidies: Where We Are and Where We're Going
Published in Darren K. Griffin, Gary L. Harton, Preimplantation Genetic Testing, 2020
Andrea Victor, Cagri Ogur, Alan Thornhill, Darren K. Griffin
Comparative genomic hybridization (CGH) is a method for analyzing copy number variations (CNVs) of a sample by comparing it to the ploidy status of a reference sample. Applying the principles of FISH, the process of CGH also utilizes DNA hybridization. However, instead of hybridizing labeled probes to a fixed sample, CGH involves fluorescently labeling sample DNA (in red) and a known euploid reference DNA (in green). These samples are then cohybridized to a normal metaphase spread to bind to their locus of origin. With the help of a fluorescent microscope and software, the differentially colored fluorescent signals are compared along the length of each chromosome for identification of chromosomal differences between the sample and reference. If the sample is euploid, the ratio of sample to reference DNA will be balanced and an equal mix of red and green will be seen across all chromosomes. Any chromosomal imbalance will be detected as a shift toward red or green for that specific chromosome.
Comparative Genomic Hybridization and Copy Number Abnormalities in Breast Cancer
Published in Brian Leyland-Jones, Pharmacogenetics of Breast Cancer, 2020
The progression of normal breast epithelial cells from a normal state toward one characterized by uncontrolled growth and metastatic behavior is caused by the deregulation of key cellular processes and signaling pathways. These alterations in normal cellular behavior are rooted in the accumulation of genomic and epigenomic lesions that impact hallmarks of cancer, such as the ability of the cell to control proliferation, undergo apoptosis, increase motility leading to invasion, and alter angiogenesis. A suite of technologies has now been developed to assess genomic and epigenomic aberrations that contribute to cancer progression. The application of these has shown that genome copy number abnormalities (CNAs) are among the most frequent genomic aberrations. Remarkably, these studies have revealed that 10% to 15% of the genes in a typical carcinoma tumor may be deregulated by recurrent genome CNAs and regions. Some of these genes influence disease progress and so may be assessed to facilitate prognosis. Others influence response to therapy and so may be assessed as predictive markers. Some of these enable oncogenic processes, on which tumors depend for survival, and so are candidate therapeutic targets. In most cases, array comparative genomic hybridization (CGH) is the method of choice for their discovery and may be used in some setting for clinical assessments of these abnormalities. Accordingly, we review here several of the current array CGH technologies available and the considerations needed when determining the technology most applicable to a given study.
Genetics
Published in Stephan Strobel, Lewis Spitz, Stephen D. Marks, Great Ormond Street Handbook of Paediatrics, 2019
Jane A. Hurst, Richard H. Scott
Until 2010 the majority of cytogenetic reports from a routine laboratory were based on analysis of G-banded chromosomes under light microscopy (Table 15.2). This allowed a visual representation of structural anomalies visible at that level of magnification. In order to diagnose submicroscopic abnormalities of less than 5 Mb, laboratories first used specific fluorescence in situ hybridisation (FISH) testing and now array comparative genomic hybridisation (CGH). Array CGH has the advantage that it detects abnormalities genome wide (i.e. at any location in the genome) in a single test.
The age-related required number of zygotes estimated from prior clinical studies of preimplantation genetic testing for aneuploidy (PGT-A)
Published in Systems Biology in Reproductive Medicine, 2023
Tasuku Mariya, Takeshi Sugimoto, Takema Kato, Toshiaki Endo, Hiroki Kurahashi
The avoidance of aneuploid embryo transfer during in vitro fertilization (IVF) has been addressed for more than 20 years in infertile couples by using preimplantation genetic testing for aneuploidy (PGT-A) (Philip et al. 1994; Vermeesch et al. 2016). After trials of various testing methods such as Fluorescent in situ hybridization (FISH), quantitative PCR, and comprehensive molecular testing with array comparative genomic hybridization (aCGH). Next-generation sequencing (NGS) is now the main analytical method of PGT-A (Kurahashi et al. 2016; Vermeesch et al. 2016). However, problems with these analyses and with the statistical methodologies when determining the efficacy of PGT-A have been reported, and caution is thus needed in interpreting the findings of these tests (Gleicher et al. 2021). In several prior Random Clinical Trials (RCTs), no differences in clinical outcomes were found between PGT-A and conventional morphological embryo sorting in certain populations (Munné et al. 2019; Yan et al. 2021). Hence, PGT-A should never be offered as a screening option, and accurate information should be provided for couples who hope to undergo this procedure.
Determining the accuracy of next generation sequencing based copy number variation analysis in Hereditary Breast and Ovarian Cancer
Published in Expert Review of Molecular Diagnostics, 2022
Nihat Bugra Agaoglu, Busra Unal, Ozlem Akgun Dogan, Payam Zolfagharian, Pari Sharifli, Aylin Karakurt, Burak Can Senay, Tugba Kizilboga, Jale Yildiz, Gizem Dinler Doganay, Levent Doganay
CNVs can be detected by several different methods like array-comparative genomic hybridization (aCGH) and single nucleotide polymorphism array (SNP-array). Both are hybridization-based methods, and despite easy application, their resolution is low, and it is not possible to determine the breakpoint regions. On the other hand, methods like real-time PCR (qPCR), fluorescent in situ hybridization (FISH) and multiplex ligation-dependent probe amplification (MLPA) can also be used to determine the CNV regions. The most significant disadvantage of these methods is that they can only be applied to the regions known to carry CNVs. Moreover, it is not cost-effective to apply these methods in genes with many exons like BRCA1 and BRCA2. Today with advanced analysis tools, in addition to SNVs, it is possible to determine structural variants like CNVs by next generation sequencing (NGS) in a single run [19,20]. The data generated by NGS can be used for prescreening of CNVs by bioinformatics algorithms in cancer predisposition genes [21–26]. There are different tools available for the interpretation of CNVs by NGS, and many of these tools use different methods like read depth, GC content and a combination of all these [24–26]. Although the results are promising, the reliability of the analysis tools are still controversial and the findings are still in need of validation for diagnostic purposes.
Advances in genetic testing and optimization of clinical management in children and adults with epilepsy
Published in Expert Review of Neurotherapeutics, 2020
Marcello Scala, Amedeo Bianchi, Francesca Bisulli, Antonietta Coppola, Maurizio Elia, Marina Trivisano, Dario Pruna, Tommaso Pippucci, Laura Canafoglia, Simona Lattanzi, Silvana Franceschetti, Carlo Nobile, Antonio Gambardella, Roberto Michelucci, Federico Zara, Pasquale Striano
The array comparative genomic hybridization (array CGH) is a molecular cytogenetic method suitable for DNA copy number variants (CNVs) analysis, such as deletions or duplications. This technique is based on the quantitative comparison between test DNA extracted from peripheral blood lymphocytes of the proband and reference DNA from healthy donors, using competitive fluorescence in situ hybridization (FISH). The resolution power is variable, ranging from 1 Mb to 100 kb, but it is at least 100-fold higher than traditional cytogenetics. Rare CNVs, some of which involve known morbid genes, contribute to approximately 10% of infantile epilepsies and 5% of epileptic encephalopathies, overall [29]. In large cohorts, array CGH studies further showed recurrent deletions in 15q13.3, 16p13.11, and 15q11.2 in patients with focal epilepsies or generalized epilepsies with ID, suggesting that these rearrangements might represent susceptibility factors for these conditions [30–32]. When epilepsy is not associated with ID or dysmorphic features/malformations, the diagnostic impact of array CGH appears to be lower [33,34].