Use of whole genome amplification and comparative genomic hybridisation to detect chromosomal copy number alterations in cell line material and tumour tissue

2004 ◽  
Vol 105 (1) ◽  
pp. 18-24 ◽  
Author(s):  
S. Hughes ◽  
G. Lim ◽  
B. Beheshti ◽  
J. Bayani ◽  
P. Marrano ◽  
...  
Keyword(s):  
Cell Line ◽  
Copy Number ◽  
Whole Genome Amplification ◽  
Tumour Tissue ◽  
Comparative Genomic Hybridisation ◽  
Comparative Genomic ◽  
Whole Genome ◽  
Copy Number Alterations ◽  
Line Material ◽  
Chromosomal Copy Number

scholarly journals Evaluation of Phi29-based whole-genome amplification for microarray-based comparative genomic hybridisation

2006 ◽  
Vol 87 (1) ◽  
pp. 75-83 ◽  
Author(s):  
Edurne Arriola ◽  
Maryou B K Lambros ◽  
Chris Jones ◽  
Tim Dexter ◽  
Alan Mackay ◽  
...  
Keyword(s):  
Whole Genome Amplification ◽  
Comparative Genomic Hybridisation ◽  
Comparative Genomic ◽  
Whole Genome ◽  
Genome Amplification

scholarly journals DNA Analysis by Restriction Enzyme (DARE) enables concurrent genomic and epigenomic characterization of single cells

2019 ◽  
Vol 47 (19) ◽  
pp. e122-e122
Author(s):  
Ramya Viswanathan ◽  
Elsie Cheruba ◽  
Lih Feng Cheow
Keyword(s):  
Dna Methylation ◽  
Restriction Enzyme ◽  
Copy Number ◽  
Dna Analysis ◽  
Whole Genome Amplification ◽  
Single Cells ◽  
Whole Genome ◽  
Copy Number Alterations ◽  
Genome Wide

Abstract Genome-wide profiling of copy number alterations and DNA methylation in single cells could enable detailed investigation into the genomic and epigenomic heterogeneity of complex cell populations. However, current methods to do this require complex sample processing and cleanup steps, lack consistency, or are biased in their genomic representation. Here, we describe a novel single-tube enzymatic method, DNA Analysis by Restriction Enzyme (DARE), to perform deterministic whole genome amplification while preserving DNA methylation information. This method was evaluated on low amounts of DNA and single cells, and provides accurate copy number aberration calling and representative DNA methylation measurement across the whole genome. Single-cell DARE is an attractive and scalable approach for concurrent genomic and epigenomic characterization of cells in a heterogeneous population.

scholarly journals SNP-based chromosomal copy number ascertainment following multiple displacement whole-genome amplification

BioTechniques ◽  
2007 ◽  
Vol 42 (1) ◽  
pp. 77-83 ◽  
Author(s):  
Jason J. Corneveaux ◽  
Michael C. Kruer ◽  
Diane Hu-Lince ◽  
Keri E. Ramsey ◽  
Victoria L. Zismann ◽  
...  
Keyword(s):  
Copy Number ◽  
Whole Genome Amplification ◽  
Whole Genome ◽  
Genome Amplification ◽  
Chromosomal Copy Number ◽  
Chromosomal Copy

High-Resolution CGH Analysis of CD34+ Cells from Lower-Risk Myelodysplastic Patients Reveals Cryptic Copy Number Alterations and Predicts Overall and Leukemia-Free Survival.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 2431-2431
Author(s):  
Daniel T. Starczynowski ◽  
Suzanne Vercauteren ◽  
Adele Telenius ◽  
Sandy Sung ◽  
Kaoru Tohyama ◽  
...  
Keyword(s):  
Risk Stratification ◽  
Copy Number ◽  
Lower Risk ◽  
Array Cgh ◽  
Comparative Genomic ◽  
Whole Genome ◽  
Copy Number Alterations ◽  
Ancillary Test ◽  
Cd34 Cells ◽  
Marrow Cells

Abstract The myelodysplastic syndromes (MDS) are a heterogeneous group of clonal hematologic malignancies characterized by peripheral cytopenias, a hypercellular marrow with ineffective hematopoiesis and a propensity to progress to acute myeloid leukemia (AML). MDS is thought to arise from a primitive (CD34+) hematopoietic cell that has acquired genetic and/or epigenetic abnormalities. Risk stratification according to the International Prognostic Scoring System (IPSS) defines patient categories that correlate with survival and the likelihood of transformation to AML. Greater than 50% of individuals diagnosed with MDS are in the lower-risk groups. The importance of cytogenetics in risk stratification has been verified in several studies, but is of limited value in patients with lower-risk subtypes because approximately 50% of these patients do not have karyotypic abnormalities detectable using standard techniques. To further our biological understanding of lower-risk subtypes of MDS, and to identify potential MDS-initiating alterations in the genome, we looked for alterations in DNA extracts from purified CD34+ marrow cells from 44 MDS lower-risk patients using a submegabase bacterial artificial chromosome (BAC) array to perform whole genome comparative genomic hybridization (CGH) analyses. These studies identified numerous cryptic structural DNA alterations that were not detectable by standard cytogenetic analysis and were also not found in 15 age-matched normal controls. Although most patients tested had a normal karyotype, 23 recurring, novel copy number alterations of a median size of 0.6 megabases were identified. These included gains at 11q24.2-qter, 17q11.2 and 17q12, and losses at 2q33.1-q33.2 and 14q12. Comparison of changes in CD34+ marrow cells with DNA from CD3+ cells isolated from the same patients showed that a recurring duplication at band 17q12 was exclusive to the CD34+ cells in 2 of 3 patients. Validation of this copy number gain at chr 17q12 by FISH confirmed duplication of the locus. In addition, whole genome array CGH analysis of CD34+ marrow cells from karyotypically normal (n = 25) and abnormal (n = 15) lower risk MDS patients revealed extensive genome alterations (involving >3 Mb) correlated with poorer overall survival, and was more frequently associated with transformation to AML as compared to IPSS stratification alone. Our studies suggest that array CGH may be useful as an ancillary test to better stratify lower-risk subtypes of MDS and, at the same time lead to the identification of early mutations that contribute to disease initiation.

Genetic Alterations Detected by High-Resolution Array Comparative Genomic Hybridization in Microdissected HRS Cells Correlate with Treatment Outcome in Classical Hodgkin Lymphoma

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 522-522
Author(s):  
Christian Steidl ◽  
Adele Telenius ◽  
Sohrab P. Shah ◽  
K-John Cheung ◽  
Lorena Barclay ◽  
...  
Keyword(s):  
Treatment Outcome ◽  
High Resolution ◽  
Disease Progression ◽  
Hodgkin Lymphoma ◽  
Copy Number ◽  
Whole Genome Amplification ◽  
Array Cgh ◽  
Whole Genome ◽  
Copy Number Alterations ◽  
Genome Amplification

Abstract INTRODUCTION: Clinical decision making in Hodgkin lymphoma (HL) is primarily based on clinical variables in part because the scarcity of the malignant Hodgkin Reed Sternberg cells (HRS cells) hampers their molecular characterization. However, more recently investigation using laser capture microdissection has allowed a more detailed analysis of these cells. The objective of this study was to detect genomic alterations in HRS cells and correlate these changes with treatment outcome. PATIENTS AND METHODS: We studied 53 patients with classical HL who were primarily treated at the BC Cancer Agency in Vancouver between 1984 and 2006. All received at least 4 cycles of polychemotherapy and stage-dependent radiotherapy if indicated. The cohort included 43 pretreatment samples and 10 biopsies taken at relapse. Treatment failure was defined as disease progression or relapse at any time (n=23), treatment success as absence of progression (n=30). Whole genome amplification (GenomePlex, Sigma) of pools from 500–1000 individually picked, microdissected HRS cells (Molecular Machines & Industries Cellcut with Nikon Eclipse TE2000-S microscope) was performed. 200 ng of amplified DNA was hybridized to 32k submegabase resolution BAC tiling arrays (SMRT) against sex-matched control-DNA. Scoring of array CGH data was performed by computational analysis using CNA-HMMer v0.1 (available at http://www.cs.ubc.ca/~sshah/acgh/) based on a Hidden Markov Model (HMM). Clustering of the 53 cases was performed using the K-medoids algorithm. Areas of amplification bias and known copy number polymorphisms were excluded. RESULTS: On average whole genome amplification generated 500-fold amplification of genomic DNA. The most frequent copy number alterations (>20% of cases) included gains of 2p14–24.3, 9p12–24.3, 12p11.21–13.33, 16p11.2–13.3, 17p11.2–13.3, 17q11.1–25.1, 19p12–13.3, 19q12–13.43, 20q11.21–13.32, 21q22.11–22.2 and losses of 1p36.31–36.33, 6q11.1–27, 7q22.1–36.3, 8p23.1–23.3, 11q22.3–25, 13q33.3–34 and Xq11.2–28. We also identified several small changes (<5 Mbs) such as loss of 1p36.32, 5q31.1 or 6q23.3 and amplification of 1q32.1, 8q24.21, 17q21.31 or 20q13.2. When comparing the different outcome groups we more frequently identified gains of chromosomal regions 12p13.31–13.33 and 16p12.1–13.3 in treatment failures (39% vs. 13%, Fisher exact p=0.05 and 43% vs. 10%, p=0.009), and losses of chromosomal regions 16q12.1–12.2 and 17p13.1–13.2 (17% vs. 0%, p=0.061 and 27% vs. 4%, p=0.061) were more frequently observed in treatment successes. We did not find significant differences of these changes between pretreatment (n=13) and relapse biopsies (n=10) of patients failing treatment. Using unsupervised analysis we identified a sample cluster of eight cases characterized by simultaneous occurrence of gains of 2p, 16p, 17p, 19q and losses of 6q. Notably, treatment failed in six of these cases. DISCUSSION: The combination of laser microdissection with subsequent WGA and high resolution array CGH provides a robust and sensitive platform for detecting chromosomal imbalances in microdissected HRS cells. We identified at high-resolution new and recurrent changes defining chromosomal regions that potentially harbor oncogenes and tumor suppressor genes crucial to the pathogenesis of HL. Furthermore, we found copy number alterations that are significantly associated with disease progression which, therefore, could serve as predictive factors for treatment outcome.

scholarly journals Whole Genome Comparative Genomic Hybridization of Ewing Sarcoma Indicates Cytoskeleton, Migration and Protein Trafficking

Proceedings ◽  
2018 ◽  
Vol 2 (25) ◽  
pp. 1547
Author(s):  
Burçin Baran ◽  
Safiye Aktaş ◽  
Hülya Tosun ◽  
Gülden Diniz ◽  
Yasemin Çakır ◽  
...  
Keyword(s):  
Comparative Genomic Hybridization ◽  
Ewing Sarcoma ◽  
Protein Trafficking ◽  
Copy Number ◽  
Rho Gtpase ◽  
Predictive Biomarkers ◽  
Comparative Genomic ◽  
Whole Genome ◽  
Copy Number Alterations ◽  
Genomic Hybridization

Ewing sarcoma is a bone and soft tissue tumor either neuroectodermal or mesenchymal originated and affecting children and adolescents. In the present study, we aimed to find out prognostic and predictive biomarkers for Ewing sarcoma. Hence, we examined the copy number alterations (and related possible genes) among ten Ewing sarcoma patient samples and possible associations with the clinical outcome. DNA extraction from formalin fixed paraffin embedded archive tissues was performed. Whole genome Comparative Genomic Hybridization (CGH) was performed by NimbleGen and recorded as single Panel Rainbow through chromosomes 1–22, X and Y. Data was interpreted by SignalMap software and genetic regions matching the deletion or amplification loci were recorded. The mean age of the patients was 8.6 years. Three of the cases were male, while seven were female. According to CGH analysis, the most common DNA copy number alterations were found in SLIT-ROBO Rho GTPase activating protein (srGAP2), RANBP2 like GRIP domain (RGPD5), nephrocystin 1 (NPHP1), GTF2I repeat domain containing 2 (GTF2IRD2), pyridoxal dependent decarboxylase domain containing 1 (PXDC1), which were found down-regulated among 7 of 10 patients. In conclusion, in our dataset the copy number alterations are mostly found in genes related with cytoskeletal elements, migration and protein trafficking among our patient group. Gene functional studies are required for better understanding the role of these genes in Ewing Sarcoma pathogenesis

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