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GenomiPhi amplification of FFPE tissue DNA for array CGH

>Problem
Low yield from GenomiPhi amplification of FFPE-extracted DNA.

Solution
Some extraction methods contain factors that will partially or completely inhibit DNA polymerases, in this case Phi29. Purification of the extracted DNA might improve results. Use of commercial kits such as Nucleon™ HT Genomic DNA Extraction Kit (GE Healthcare) will reduce or eliminate inhibitory factors and can be a useful substitute.

Problem
No signal observed from microarray hybridization.

Solution
Verify that Cy3 and Cy5 probe labeling was successful by analyzing sample on gel. Check for PCR amplification of select human genes (300–900 bp) from FFPE-extracted DNA and amplified DNA to make sure DNA is of high purity.

Problem
Log2 ratios distorted.

Solution
Use test and reference samples amplified under similar conditions. Make sure repetitive sequences are repressed by pre-hybridizing the arrays with human Cot-1 DNA™ (Invitrogen) and denatured herring sperm DNA.


References
1. Salman, M. et al. Will the new cytogenetics replace the old cytogenetics? Clin. Genet. 66, 265–275 (2004).

2. Kallioniemi, A. et al. Comparative genomic hybridization for molecular cytogenetic analysis of solid tumors. Science 258, 818–821 (1992).

3. Fiegler, H. et al. DNA microarrays for comparative genomic hybridization based on DOP-PCR amplification of BAC and PAC clones. Genes Chromosomes Cancer 36, 361–374 (2003).

4. Mamone, J. T. A method for representatively amplifying genomic DNA. Genomic/Proteomic
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