Array Comparative Genomic Hybridization: Protocols and by Diponkar Banerjee (auth.), Diponkar Banerjee, Sohrab P. Shah

By Diponkar Banerjee (auth.), Diponkar Banerjee, Sohrab P. Shah (eds.)

The earliest descriptions of human chromosomes initiated the genomics revolution that's now upon us. Array Comparative Genomic Hybridization: Protocols and Applicationsexplores the scope of what's now attainable so far as documenting abnormalities linked to numerous human cancers. whereas the know-how for interrogating the human genome keeps to adapt, refinement of comparative genomic hybridization (CGH) utilizing array CGH and similar applied sciences have supplied huge, immense perception into human cancers at a reasonable scale. Written within the profitable Methods in Molecular Biology™ sequence layout, chapters comprise introductions to their respective subject matters, lists of the mandatory fabrics and reagents, step by step, quite simply reproducible protocols, and notes on troubleshooting and warding off recognized pitfalls.

Authoritative and simply available, Array Comparative Genomic Hybridization: Protocols and Applicationsprovides researchers with well-honed methodologies to benefit those strategies for his or her personal use in learn or medical diagnostic laboratories.

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1). Snap-frozen biopsies are an excellent source of high quality DNA, usually in sufficient amounts for aCGH. An important cancer sample issue is tumor heterogeneity. Tumorigenesis is a micro-evolution in which variants of cells clonally expand within a tissue. This dynamic process results in genetic diversity within tumors and marked changes in tissue architecture. Thus tumor samples typically contain various proportions of tumor cells and normal cells as well as components of stroma and infiltrating lymphocytes (11).

To extract and process the data, the FE software first creates and aligns a grid to the image, identifies image spots, recognizes and rejects outlier pixels and then flags outlier features. Finally, signal and background biases are computed, calculation of signal log ratios is performed, and errors are estimated. The log ratio of fluorescence is the final readout of each spot on the array. 2 Ultradense Array CGH and Discovery of Micro-Copy Number… 27 Explicit steps to run FE are found in the user guide (20) and briefly described in Agilent’s Feature Extraction software tutorial (21).

Elute with 30 μl of nanopure (Milli-Q) autoclaved water (allow 5 min for elution). 10. Centrifuge full speed for 1 min. 11. Elute again with another 30 μl of water. 12. Centrifuge at full speed (13,000 rpm (16,000 × g)) for 1 min. 13. Use NanoDrop to determine DNA concentration and purity (see Note 13). 14. To obtain a more precise measurement of DNA concentration in the samples use PicoGreen ds DNA Assay Kit (Invitrogen/ Molecular Probes #P11496). 15. If concentration of DNA is less than 10ng/µl (PicoGreen values), concentrate the samples using SpeedVac, (see Note 14).

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