Summary
Comparative genomic hybridization (CGH) is arguably the most significant technical development in the molecular cytogenetics era, and has contributed considerably to our further understanding of the cancer genome. In essence, DNA from a cancer specimen (test DNA) labeled with the fluorescence reporter molecule (or fluorochrome) is hybridized to a target genome in the presence of a differentially labeled control DNA (reference DNA). The two DNA populations compete for hybridization sites on normal metaphase chromosomes, so that the resulting fluorescence ratio is a reflection of the copy number change in the test sample. The copy number changes are mapped to their position on the chromosome template. Over recent years, the chromosomal template has been largely superseded by microarray formats (aCGH), in which changes in copy number can be mapped to the genome sequence at a high resolution. This advance allows the genome to be studied at an unbridled resolution and at a high-throughput, whilst posing several technical, statistical and interpretive challenges. It is the aim of this chapter to introduce the fundamental concepts of aCGH and to provide an overview of the steps involved in a successful aCGH processing. The materials required for BAC and oligonucleotide aCGH are included, with detailed methods and a range of refinements to improve the success rate and quality of aCGH data.
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References
Kallioniemi, A., et al. Comparative genomic hybridisation for molecular genetic analysis of solid tumours. Science 258(5083),818–821 (1992).
Solinas-Toldo, S., et al. Matrix-based comparative genomic hybridization: biochips to screen for genomic imbalances. Genes Chromosomes Cancer 20, 399–407 (1997).
Pinkel, D., et al. High resolution analysis of DNA copy number variation using comparative genomic hybridization to microarrays. Nat Genet 20, 207–211 (1998).
Snijders, A.M., et al. Assembly of microarrays for genome-wide measurement of DNA copy number. Nat Genet 29, 263–264 (2001).
Fiegler, H., et al. DNA microarrays for comparative genomic hybridization based on DOP-PCR amplification of BAC and PAC clones. Genes Chromosomes Cancer 37, 223 (2003).
Smirnov, D.A., Burdick, J.T., Morley, M. & Cheung, V.G. Method for manufacturing whole-genome microarrays by rolling circle amplification. Genes Chromosomes Cancer 40, 72–77 (2004).
Ishkanian, A.S., et al. A tiling resolution DNA microarray with complete coverage of the human genome. Nat Genet 36, 299–303 (2004).
Krzywinski, M., et al. A set of BAC clones spanning the human genome. Nucleic Acids Res 32, 3651–3660 (2004).
Pollack, J.R., et al. Genome-wide analysis of DNA copy-number changes using cDNA microarrays. Nat Genet 23, 41–46 (1999).
Dhami, P., et al. Exon array CGH: detection of copy-number changes at the resolution of individual exons in the human genome. Am J Hum Genet 76, 750–762 (2005).
Brennan, C., et al. High-resolution global profiling of genomic alterations with long oligonucleotide microarray. Cancer Res 64, 4744–4748 (2004).
Callagy, G., et al. Identification and validation of prognostic markers in breast cancer with the complementary use of array-CGH and tissue microarrays. J Pathol 205, 388–396 (2005).
Barrett, M.T., et al. Comparative genomic hybridization using oligonucleotide microarrays and total genomic DNA. Proc Natl Acad Sci USA 101, 17765–17770 (2004).
Schouten, J.P., et al. Relative quantification of 40 nucleic acid sequences by multiplex ligation-dependent probe amplification. Nucleic Acids Res 30, e57 (2002).
Daser, A., et al. Interrogation of genomes by molecular copy-number counting (MCC). Nat Methods 3, 447–453 (2006).
Strefford, J.C., et al. Genome complexity in acute lymphoblastic leukaemia is revealed by array-based comparative genomic hybridization. Oncogene 26, 4306–4318 (2007).
Paulsson, K., et al. Identification of cryptic aberrations and characterization of translocation breakpoints using array CGH in high hyperdiploid childhood acute lymphoblastic leukemia. Leukemia 20, 2002–2007 (2006).
Moorman, A.V., et al. Prognosis of children with acute lymphoblastic leukaemia (ALL) and intrachromosomal amplification of chromosome 21 (iAMP21). Blood 109, 2327–2330 (2007).
Strefford, J.C., et al. Complex genomic alterations and gene expression in acute lymphoblastic leukemia with intrachromosomal amplification of chromosome 21. Proc Natl Acad Sci USA 103, 8167–8172 (2006).
van Vlierberghe, P., et al. A new recurrent 9q34 duplication in pediatric T-cell acute lymphoblastic leukemia. Leukemia 20, 1245–1253 (2006).
van Vlierberghe, P., et al. The cryptic chromosomal deletion, del(11)(p12p13), as a new activation mechanism of LMO2 in pediatric T-cell acute lymphoblastic leukemia. Blood 108, 3520–3529 (2006).
Barber, K.E., et al. Amplification of the ABL gene in T-cell acute lymphoblastic leukemia. Leukemia 18, 1153–1156 (2004).
Graux, C., et al. Fusion of NUP214 to ABL1 on amplified episomes in T-cell acute lymphoblastic leukemia. Nat Genet 36, 1084–1089 (2004).
Jong, K., Marchiori, E., Meijer, G., Vaart, A.V. & Ylstra, B. Breakpoint identification and smoothing of array comparative genomic hybridization data. Bioinformatics 20, 3636–3637 (2004).
Hupe, P., Stransky, N., Thiery, J.P., Radvanyi, F. & Barillot, E. Analysis of array CGH data: from signal ratio to gain and loss of DNA regions. Bioinformatics 20, 3413–3422 (2004).
Eilers, P.H. & de Menezes, R.X. Quantile smoothing of array CGH data. Bioinformatics 21, 1146–1153 (2005).
Acknowledgements
The authors are grateful to Professor Christine Harrison for critical reading of the manuscript. This work was funded by Leukaemia Research, United Kingdom.
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Strefford, J., Parker, H. (2009). Array-Based Comparative Genomic Hybridization as a Tool for Analyzing the Leukemia Genome. In: Eric So, C.W. (eds) Leukemia. Methods in Molecular Biology™, vol 538. Humana Press. https://doi.org/10.1007/978-1-59745-418-6_8
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DOI: https://doi.org/10.1007/978-1-59745-418-6_8
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