Detailed chromosomal and molecular genetic analysis of single cells by whole genome amplification and comparative genomic hybridisation

Wells, Dagan; Sherlock, Jon; Handyside, Alan H.; Delhanty, Joy D. A.

doi:10.1093/nar/27.4.1214

Cited by 283 publications

(152 citation statements)

References 6 publications

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“…Amplification and fluorescent labeling of the DNA from microdissected tissue was carried out by DOP-PCR in two rounds as previously published (Wells et al, 1999), using 2 to 5 l of the total volume of extracted DNA. Normal male metaphase spreads (Vysis UK Ltd, Richmond, England) were denatured at 75°C for 5 minutes in 70% formamide, 2ϫ SSC, and dehydrated through a series of alcohols.…”

Section: Comparative Genomic Hybridization Analysismentioning

confidence: 99%

Comparative Genomic Hybridization Analysis of Myoepithelial Carcinoma of the Breast

Jones

Foschini

Chaggar³

et al. 2000

Laboratory Investigation

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SUMMARY:Although there seems to be a common stem cell for the two epithelial cell types in the breast, the vast majority of breast cancers exhibit a luminal phenotype. Pure myoepithelial carcinomas are rare. We report our findings of genetic alterations in these tumors. We have analyzed 10 cases of pure myoepithelial cell carcinomas using laser capture microdissection and comparative genomic hybridization. The mean number of changes was 2.1 (range 0 -4), compared with a mean of 8.6 (range 3.6 -13.8) in unselected ductal carcinomas. Common alterations included loss at 16q (3/10 cases), 17p (3/10), 11q (2/10), and 16p (2/10), regions also commonly deleted in ductal carcinomas. The single case in which both pure myoepithelial carcinoma and invasive ductal carcinoma was present showed 2 alterations in the myoepithelial tumor (losses at 17p and 17q), whereas the invasive ductal component showed 15 alterations (5 gains and 9 losses), including loss at 17p. The sharing of 17p loss in myoepithelial and ductal carcinoma is consistent with a common stem cell model in the breast. The relatively few genetic alterations in otherwise aggressive neoplasms suggests that myoepithelial tumors may be a good model for the delineation of genes important in breast tumorigenesis. (Lab Invest 2000, 80:831-836).

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Section: Comparative Genomic Hybridization Analysismentioning

confidence: 99%

Comparative Genomic Hybridization Analysis of Myoepithelial Carcinoma of the Breast

Jones

Foschini

Chaggar³

et al. 2000

Laboratory Investigation

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“…In addition, the use of wholegenome amplification (WGA) techniques to amplify extremely small amounts of genomic DNA would enable analysis directly on amniotic fluid (AF) cells or chorionic villus samples (CVS) without culturing, leading to more rapid results without the added time or cost of cell culture. 12,13 There are several reports of the use of array-CGH for multiplex diagnosis of cytogenetic abnormalities using blood samples and at least one report of the application of array-CGH to previously studied, archived prenatal samples, but to date, there has been no experience with the use of this technique to monitor and report on current pregnancies. 11,14 -19 We designed a targeted microarray of 366 large genomic clones covering the 41 clinically relevant subtelomeric regions as well as genomic positions corresponding to 55 different genomic disorders, validated its use for molecular cytogenetic diagnosis on samples with known abnormalities, and completed over 1,500 clinical cases on referred blood samples.…”

mentioning

confidence: 99%

Prenatal diagnosis of chromosomal abnormalities using array-based comparative genomic hybridization

Sahoo

Cheung

Ward

et al. 2006

Genetics in Medicine

156

131

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Purpose: This study was designed to evaluate the feasibility of using a targeted array-CGH strategy for prenatal diagnosis of genomic imbalances in a clinical setting of current pregnancies. Methods: Women undergoing prenatal diagnosis were counseled and offered array-CGH (BCM V4.0) in addition to routine chromosome analysis.Array-CGH was performed with DNA directly from amniotic fluid cells with whole genome amplification, on chorionic villus samples with amplification as necessary, and on cultured cells without amplification. Results: Ninety-eight pregnancies (56 amniotic fluid and 42 CVS specimens) were studied with complete concordance between karyotype and array results, including 5 positive cases with chromosomal abnormalities. There was complete concordance of array results for direct and cultured cell analysis in 57 cases tested by both methods. In 12 cases, the array detected copy number variation requiring testing of parental samples for optimal interpretation. Array-CGH results were available in an average of 6 and 16 days for direct and cultured cells, respectively. Patient acceptance of array-CGH testing was 74%. Conclusion: This study demonstrates the feasibility of using array-CGH for prenatal diagnosis, including reliance on direct analysis without culturing cells. Use of array-CGH should increase the detection of abnormalities relative to the risk, and is an option for an enhanced level of screening for chromosomal abnormalities in high risk pregnancies. Genet Med 2006:8(11):719-727.

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“…[1][2][3] However, it is difficult to verify the results of single-cell CGH as the respective cell is destroyed during the procedure and therefore unavailable for further experiments.…”

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confidence: 99%

Sequential application of interphase-FISH and CGH to single cells

Langer

Geigl

Gangnus

et al. 2005

Laboratory Investigation

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A comprehensive genomic analysis of single cells is needed for numerous scenarios in tumor genetics, clinical diagnostics and forensic application. PCR protocols were developed which allow an unbiased amplification of the whole genome of a single cell for subsequent analyses by comparative genomic hybridization (CGH). However, verification of single-cell CGH results has been impossible as the procedure naturally involves the destruction of the respective cell. Here we show that the genome of individual cells can be analyzed by two different single cell techniques applied sequentially to the same cell. In a first step, interphase fluorescence in situ hybridization (FISH) is applied. After evaluation of the interphase-FISH signals, cells of interest can be selected for a further analysis. Single cells are collected by laser microdissection, the DNA is amplified by linker-adaptor PCR and subjected to CGH-analysis. This strategy offers new opportunities for a sophisticated selection of cells based on interphase-FISH signals. Furthermore, the sequential application of two different single-cell approaches to the same single-cell represents the only option to control and verify the single-cell CGH results. We demonstrate the feasibility of this approach with a series of experiments including cells from pre-and postnatal diagnostics, for example, cells with trisomies 13, 18, or 21, respectively, leukemia and tumor cells and tissue sections.

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Detailed chromosomal and molecular genetic analysis of single cells by whole genome amplification and comparative genomic hybridisation

Cited by 283 publications

References 6 publications

Comparative Genomic Hybridization Analysis of Myoepithelial Carcinoma of the Breast

Comparative Genomic Hybridization Analysis of Myoepithelial Carcinoma of the Breast

Prenatal diagnosis of chromosomal abnormalities using array-based comparative genomic hybridization

Sequential application of interphase-FISH and CGH to single cells

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