Article

Guidelines for molecular karyotyping in constitutional genetic diagnosis

Center for Human Genetics, University Hospital Gasthuisberg, Leuven, Belgium.
European Journal of HumanGenetics (Impact Factor: 4.23). 12/2007; 15(11):1105-14. DOI: 10.1038/sj.ejhg.5201896
Source: PubMed

ABSTRACT Array-based whole genome investigation or molecular karyotyping enables the genome-wide detection of submicroscopic imbalances. Proof-of-principle experiments have demonstrated that molecular karyotyping outperforms conventional karyotyping with regard to detection of chromosomal imbalances. This article identifies areas for which the technology seems matured and areas that require more investigations. Molecular karyotyping should be part of the genetic diagnostic work-up of patients with developmental disorders. For the implementation of the technique for other constitutional indications and in prenatal diagnosis, more research is appropriate. Also, the article aims to provide best practice guidelines for the application of array comparative genomic hybridisation to ensure both technical and clinical quality criteria that will optimise and standardise results and reports in diagnostic laboratories. In short, both the specificity and the sensitivity of the arrays should be evaluated in every laboratory offering the diagnostic test. Internal and external quality control programmes are urgently needed to evaluate and standardise the test results between laboratories.

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    • "When interpreting and classifying CNVs, it is essential to distinguish gains from losses because the potential clinical consequences might significantly differ. Furthermore, it is essential to compare gains with gains and losses with losses (Vermeesch et al., 2007; Conrad et al., 2010; Vermeesch et al., 2012). de Leeuw et al. (2012) summarized the characteristics of the most commonly used Internet databases and resources and proposed a general interpretation strategy that can be used for comparative hybridization, comparative intensity and genotype-based array data. "
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    ABSTRACT: The field of cytogenetics has focused on studying the number, structure, function and origin of chromosomal abnormalities and the evolution of chromosomes. The development of fluorescent molecules that either directly or via an intermediate molecule bind to DNA has led to the development of fluorescent in situ hybridization (FISH), a technology linking cytogenetics to molecular genetics. This technique has a wide range of applications that increased the dimension of chromosome analysis. The field of cytogenetics is particularly important for medical diagnostics and research as well as for gene ordering and mapping. Furthermore, the increased application of molecular biology techniques, such as array-based technologies, has led to improved resolution, extending the recognized range of microdeletion/microduplication syndromes and genomic disorders. In adopting these newly expanded methods, cytogeneticists have used a range of technologies to study the association between visible chromosome rearrangements and defects at the single nucleotide level. Overall, molecular cytogenetic techniques offer a remarkable number of potential applications, ranging from physical mapping to clinical and evolutionary studies, making a powerful and informative complement to other molecular and genomic approaches. This manuscript does not present a detailed history of the development of molecular cytogenetics; however, references to historical reviews and experiments have been provided whenever possible. Herein, the basic principles of molecular cytogenetics, the technologies used to identify chromosomal rearrangements and copy number changes, and the applications for cytogenetics in biomedical diagnosis and research are presented and discussed.
    Genetics and Molecular Biology 03/2014; 37(1 Suppl):194-209. DOI:10.1590/S1415-47572014000200006 · 0.88 Impact Factor
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    • "Because microarray data is dependent on array platform, the quality of DNA and software algorithms, it is important that individual laboratories develop their own protocols for assessing mosaicism. This has been recommended by both the American College of Medical Genetics and the European best practices guidelines for constitutional microarray testing [17,18]. As a prerequisite for assessing the accuracy of arrays for the identification of embryonic mosaicism, we first determined the limits of detection of mosaicism by performing reconstitution experiments using DNA mixed from normal and aneuploid samples and analyzing CMA results for mosaicism detection using BlueGnome’s 24sure array and data generated by BlueGnome’s BlueFuse Multi software. "
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    ABSTRACT: Most previous studies of chromosomal mosaicism in IVF embryos were performed by fluorescence in situ hybridization (FISH) methods. While there are reports implicating chromosome aneuploidy in implantation failure following transfer and pregnancy loss by spontaneous miscarriage, the significance of mosaicism for the developmental potential of growing embryos is unknown. However, the low prevalence of chromosomal mosaicism in chorionic villus sampling and amniotic fluid specimens suggests the presence of selection against mosaic embryos for implantation and early pregnancy. The absence of evidence for selective allocation of abnormal cells to the trophectoderm (TE) of mosaic blastocysts permits these cells to be a good proxy for embryonic mosaicism detection by chromosomal microarrays (CMA). The purpose of this study was to establish the limits of detection and the prevalence of chromosome mosaicism in day 5/6 human embryos using CMA with TE biopsies. From reconstitution experiments we established log2 ratio thresholds for mosaicism detection. These studies indicated that chromosomal mosaicism at levels as low as between 25-37% can be consistently identified. Follow-up studies by FISH on non-transferred abnormal embryos confirmed the diagnostic accuracy of CMA testing. The number of cells in a TE biopsy can influence mosaicism detection. Chromosomal microarrays can detect mosaicism in TE biopsies when present at levels as low as between 25-37% and the prevalence of day 5/6 blastocysts which were mosaic and had no other abnormalities reached 15% among a cohort of 551 embryos examined. Validated protocols for establishing detection thresholds for mosaicism are important to reduce the likelihood of transferring abnormal embryos.
    Molecular Cytogenetics 02/2014; 7(1):18. DOI:10.1186/1755-8166-7-18 · 2.66 Impact Factor
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    • "In eight (4.8%) samples a CNV inherited from a healthy parent was detected. Although not all inherited CNVs can always be classified as benign and without clinical relevance with certainty, they are, dependent on the size and the type of CNV, less likely to directly lead to a clinical phenotype: small CNVs (< 0.1 Mb) and gains are less likely to be pathogenic than large CNVs (>1 Mb) and losses, respectively [33,34]. All inherited CNVs in this study were gains ranging in size from ~250 - 1,200 kb, mostly maternally inherited and most likely benign. "
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    Molecular Cytogenetics 01/2014; 7(1):6. DOI:10.1186/1755-8166-7-6 · 2.66 Impact Factor
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