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GenomiPhi amplification of FFPE tissue DNA for array CGH

iseased tissues (2, 3). CGH experiments can be performed with DNA purified from cell culture or generated using amplification methods for fluorescent probe labeling, particularly when sample amount is limited.

Profiling DNA copy-number changes in FFPE DNA has the potential to reveal genetic information on particular regions of genomes that might be altered in clinically relevant samples. However, FFPE samples pose a unique challenge to CGH analysis because DNA isolated from them is degraded and requires amplification before array analysis.

Whole-genome amplification (WGA) methods are used to yield sufficient template for probe labeling and array hybridization. A common such method is degenerate oligonucleotide–primed PCR (DOP-PCR).

In contrast to DOP-PCR, GenomiPhi™ DNA Amplification Kit provides WGA using a simple isothermal method (4). This method employs the unique biochemical properties of Phi29 DNA polymerase, a highly processive enzyme with excellent strand displacement and proofreading ability, to amplify DNA.

In this application note, we show array CGH data on DNA isolated from human liver FFPE slides, comparing DOP-PCR with GenomiPhi DNA Amplification Kit. The amplification methods are validated by comparing microarray hybridization data on human BAC arrays. The results show that the GenomiPhi method provides robust whole-genome amplification of FFPE material, with array CGH data quality similar to the DOP-PCR method.


Materials
Products used
GenomiPhi DNA Amplification Kit, 25-6600-01
100 reactions

Cy3-dCTP PA53021

Cy5-dCTP PA55021

Typhoon™
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Source:


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