( nomiPhi (including modified and unmodif...)GenomiPhi: protocol for BAC amplification and fabrication of slides for array CGH

nomiPhi (including modified and unmodified nucleotides) and DOP-PCR methods: The DNA concentration was adjusted to 100 ng/µl with water. An equal volume of 2×printing buffer (200 mM sodium phosphate, pH 8.5) was added to the DNA sample, mixed thoroughly and spotted onto CodeLink Activated and UltraGAPS slides using the Piezorray™ spotter to generate eight replicate spots at a relative humidity of 42%. UV cross-linking (3000 µJoules) was used to immobilize the DNA onto the UltraGAPS slides. Coupling of DNA to CodeLink slides was achieved by incubating the slides in a saturated NaCl chamber for 48 h. Refer to the CodeLink user guide (GE Healthcare) for further processing of the CodeLink slides.

Conclusions
The GenomiPhi DNA Amplification Kit can be used successfully to generate BAC arrays. Compared to DOPPCR, the GenomiPhi method provides a simple, cheap and easy to use protocol. Analysis of microarray data showed that both methods produced similar DNA yields, sensitivity and performance. We also showed that BAC DNA generated by the GenomiPhi method detected all 8p12 chromosomal amplifications that were also detected by the DOP-PCR method in an MDA-MB-134 breast cancer line.

References
1. Mamone, J. T. A method for representatively amplifying genomic DNA. Genomic/Proteomic Technology April/May 2003. [Online.] http://www.amershambiosciences.com/applic/upp00738.nsf/vLookup Doc/253872879-B500/$file/GPT_Mamone_Reprint.pdf

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

3. Solinas-Toldo, S. et al. Matrix-based comparative genomic hybridization: biochips to screen for genomic imbalances. Genes Chromosomes Cancer 20, 399–407 (1997).

4. Fiegler, H. et al.
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