Cytogenetic analysis of human blastocysts with the use of FISH, CGH and aCGH: scientific data and technical evaluation

Fragouli, Elpida; Alfarawati, S; Daphnis, D; Goodall, N.-N.; Mania, Anastasia; Griffiths, Tracey; Gordon, Alexander; Wells, Dagan

doi:10.1093/humrep/deq344

Cited by 258 publications

(177 citation statements)

References 30 publications

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“…The Sureplex WGA procedure consists of three steps: extraction and fragmentation of the genomic DNA of the isolated cells to fragments in the range of a few hundred basepairs, ligation of specific adaptor sequences to both ends of the fragments and a subsequent amplification reaction by PCR using the flanking universal priming sites contained in the adaptors. This system is widely used in the clinical setting for pre-implantation genetic diagnosis starting from single-cell material 34 and, although it only amplifies a representation of the genome, has a relatively low allele-dropout rate and amplification bias 22,23 .…”

Section: Methodsmentioning

confidence: 99%

Low-grade chromosomal mosaicism in human somatic and embryonic stem cell populations

et al. 2014

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Current knowledge on chromosomal mosaicism in human cell cultures is mostly based on cytogenetic banding methods. The recent development of high-resolution full-genome analysis methods applicable to single cells is providing new insights into genetic and cellular diversity. Here we study the genetic content of 92 individual human cells, including fibroblasts, amniocytes and embryonic stem cells (hESCs), using single-cell array-based comparative genomic hybridization (aCGH). We find that human somatic and embryonic stem cell cultures show significant fractions of cells carrying unique megabase-scale chromosomal abnormalities, forming genetic mosaics that could not have been detected by conventional cytogenetic methods. These findings are confirmed by studying seven clonal hESC sub-lines by aCGH. Furthermore, fluorescent in situ hybridisation reveals an increased instability of the subtelomeric regions in hESC as compared to somatic cells. This genetic heterogeneity may have an impact on experimental results and, in the case of hESC, on their potential clinical use.

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Section: Methodsmentioning

confidence: 99%

Low-grade chromosomal mosaicism in human somatic and embryonic stem cell populations

et al. 2014

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“…Like cleavage stage biopsy, mosaicism can also yield misleading results when using trophectoderm biopsy for sampling. This problem is partially mitigated by the developmental stage of the embryo (some abnormal embryos may not survive 5 d in culture, Adler et al 2014) and the ability to access and analyze several trophectoderm cells (Fragouli et al 2011;Capalbo et al 2013).…”

Section: Genetic Considerations In Recurrent Pregnancy Lossmentioning

confidence: 99%

Genetic Considerations in Recurrent Pregnancy Loss

Hyde

Schust

2015

Cold Spring Harbor Perspectives in Medicine

118

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“…Still, they are much more sensitive than cytogenetic assays, based on fluorescence in situ hybridization (FISH), which have a resolution of no more than 10Mb[34], but offer the advantage that low-abundant chromosomal mutations can be detected in single cells. More recently, single cell assays have also emerged for aCGH based on whole genome amplification[35–37]. However, while useful in their own right, aCGH and FISH are incapable of detecting small mutations, which comprise the majority of the mutational landscape.…”

Section: Introductionmentioning

confidence: 99%

Direct mutation analysis by high-throughput sequencing: From germline to low-abundant, somatic variants

Gundry

Vijg

2012

Mutation Research/Fundamental and Molecular Mechanisms of Mutag

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DNA mutations are the source of genetic variation within populations. The majority of mutations with observable effects are deleterious. In humans mutations in the germ line can cause genetic disease. In somatic cells multiple rounds of mutations and selection lead to cancer. The study of genetic variation has progressed rapidly since the completion of the draft sequence of the human genome. Recent advances in sequencing technology, most importantly the introduction of massively parallel sequencing (MPS), have resulted in more than a hundred-fold reduction in the time and cost required for sequencing nucleic acids. These improvements have greatly expanded the use of sequencing as a practical tool for mutation analysis. While in the past the high cost of sequencing limited mutation analysis to selectable markers or small forward mutation targets assumed to be representative for the genome overall, current platforms allow whole genome sequencing for less than $5,000. This has already given rise to direct estimates of germline mutation rates in multiple organisms including humans by comparing whole genome sequences between parents and offspring. Here we present a brief history of the field of mutation research, with a focus on classical tools for the measurement of mutation rates. We then review MPS, how it is currently applied and the new insight into human and animal mutation frequencies and spectra that has been obtained from whole genome sequencing. While great progress has been made, we note that the single most important limitation of current MPS approaches for mutation analysis is the inability to address low-abundance mutations that turn somatic tissues into mosaics of cells. Such mutations are at the basis of intra-tumor heterogeneity, with important implications for clinical diagnosis, and could also contribute to somatic diseases other than cancer, including aging. Some possible approaches to gain access to low-abundance mutations are discussed, with a brief overview of new sequencing platforms that are currently waiting in the wings to advance this exploding field even further.

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Cytogenetic analysis of human blastocysts with the use of FISH, CGH and aCGH: scientific data and technical evaluation

Cited by 258 publications

References 30 publications

Low-grade chromosomal mosaicism in human somatic and embryonic stem cell populations

Low-grade chromosomal mosaicism in human somatic and embryonic stem cell populations

Genetic Considerations in Recurrent Pregnancy Loss

Direct mutation analysis by high-throughput sequencing: From germline to low-abundant, somatic variants

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