Oligonucleotide array CGH was performed using the Agilent Human Genome CGH Microarray Kit 44 K®. This microarray consisted of more than 44,000 oligonucleotide probes that spanned both coding and non-coding regions. The coverage of the human genome was made with an average spatial resolution of 75,000 pair bases.
The patient’s DNA as well as a reference DNA was fragmented by heat at 95 °C for 20 min. Each fragmented DNA product was labeled by random priming using either ULS5 or ULS3. After column-purification, probes were denatured and pre-annealed with 5 μg of human Cot-1 DNA, 10 μl of CGH Blocking agent and 55 μl of hybridization buffer. Hybridization was performed at 65 °C during 24 h. The microarray was washed, scanned and analyzed with Agilent Feature Extraction® 9.1 software. Results were interpreted using DNA analytics® 4.5 software. Only imbalances involving three or more adjacent probes were held. The identification of probes with a significant gain or loss was based on the log2 ratio plot deviation from 0 with cutoff values of 0.5 to 1, and − 0.5 to − 1, respectively.
The patient’s DNA as well as a reference DNA was fragmented by heat at 95 °C for 20 min. Each fragmented DNA product was labeled by random priming using either ULS5 or ULS3. After column-purification, probes were denatured and pre-annealed with 5 μg of human Cot-1 DNA, 10 μl of CGH Blocking agent and 55 μl of hybridization buffer. Hybridization was performed at 65 °C during 24 h. The microarray was washed, scanned and analyzed with Agilent Feature Extraction® 9.1 software. Results were interpreted using DNA analytics® 4.5 software. Only imbalances involving three or more adjacent probes were held. The identification of probes with a significant gain or loss was based on the log2 ratio plot deviation from 0 with cutoff values of 0.5 to 1, and − 0.5 to − 1, respectively.