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Digital karyotyping

Abstract

Detection of copy number variation in the human genome is important for identifying naturally occurring copy number polymorphisms as well as alterations that underlie various human diseases, including cancer. Digital karyotyping uses short sequence tags derived from specific genomic loci to provide a quantitative and high-resolution view of copy number changes on a genome-wide scale. Genomic tags are obtained using a combination of enzymatic digests and isolation of short DNA sequences. Individual tags are linked into ditags, concatenated, cloned and sequenced. Tags are matched to reference genome sequences and digital enumeration of groups of neighboring tags provides quantitative copy number information along each chromosome. Digital karyotyping libraries can be generated in about a week, and library sequencing and data analysis require several additional weeks.

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Figure 1: Schematic outline of the DK protocol.
Figure 2: Gel electrophoresis to verify the integrity of genomic DNA.
Figure 3: Optimization of PCR amplification of ditags.
Figure 4: Representative visual output from CGAP DKView.

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Acknowledgements

The authors are supported by the Ludwig Trust, the Pew Charitable Trusts, NIH grants CA121113, CA57345 and CA62924, and the NCI Division of Cancer Prevention contract HHSN261200433002C.

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Correspondence to Victor E Velculescu.

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Competing interests

Under a licensing agreement between Genzyme and the Johns Hopkins University, Dr. Velculescu is entitled to a share of royalty received by the University on sales of products described in this article. Dr. Velculescu and the University own Genzyme stock, which is subject to certain restrictions under University policy. The terms of this arrangement are being managed by the Johns Hopkins University in accordance with its conflict of interest policies.

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Leary, R., Cummins, J., Wang, TL. et al. Digital karyotyping. Nat Protoc 2, 1973–1986 (2007). https://doi.org/10.1038/nprot.2007.276

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