Cgh analytics software v3

DNA extracted from the cell line was compared to a reference sample for standard two-color aCGH. Reference DNA was purchased from Promega (Madison, WI, USA). Test samples were labeled using Cy5 and Reference DNA was labeled using Cy3. Agilent 244K human genome oligonucleotide aCGH (G4411B) was used for aCGH analysis following the manufacturer’s instructions (Oligonucleotide Array-Based CGH for Genomic DNA Analysis protocol version 3 (Agilent Technologies, Palo Alto, CA, USA). Images were captured using an Agilent scanner and quantified using Feature Extraction software v9.0 (Agilent Technologies, Palo Alto, CA, USA). CGH analytics software v3.4 (Agilent Technologies, Palo Alto, CA, USA) was subsequently used for data normalization, quality evaluation, and data visualization. Copy number aberration was indicated using the ADM-2 (Aberration Detection Method 2) algorithm. Probe positions were mapped to GRCh38.

DNA extracted from the cell line was compared to a reference sample for standard two-color aCGH. Reference DNA was purchased from Promega (Madison, WI, USA). Test samples were labeled using Cy5 and Reference DNA was labeled using Cy3. Agilent 244K human genome oligonucleotide aCGH (G4411B) was used for aCGH analysis following the manufacturer’s instructions (Oligonucleotide Array-Based CGH for Genomic DNA Analysis protocol version 3 (Agilent Technologies, Palo Alto, CA, USA). Images were captured using an Agilent scanner and quantified using Feature Extraction software v9.0 (Agilent Technologies, Palo Alto, CA, USA). CGH analytics software v3.4 (Agilent Technologies, Palo Alto, CA, USA) was subsequently used for data normalization, quality evaluation, and data visualization. Copy number aberration was indicated using the ADM-2 (Aberration Detection Method 2) algorithm. Probe positions were mapped to GRCh38.

Array-based comparative genomic hybridization (array-CGH) was performed by Miltenyi Biotec (Bergisch Gladbach, Germany) while using an Agilent Human Genome CGH Microarray 244K (Agilent Technologies, Waldbronn, Germany) consisting of 244,000 in situ synthesized 60-mer oligonucleotides spanning the entire human genome with an average probe spacing of 6.5 kb.
Genomic DNA of the SRH cell line and a female reference DNA (Promega, Mannheim, Germany) for control were labeled with Cy5-dCTP and Cy3-dCTP, respectively, and then hybridized to an oligonucleotide microarray, according to the Agilent oligonucleotide array-based CGH for genomic DNA analysis protocol v5.0 while using the Agilent Oligo aCGH Hybridization Kit. The fluorescence signals of the hybridized microarray were detected using Agilent’s DNA microarray scanner. The Agilent Feature Extraction software was used in order to read out and process the microarray image file. Further analysis and the visualization of the hybridization result were performed with the Agilent CGH Analytics software v3.4 with the following aberration filter settings: a minimum number of probes present in an aberrant region = 2; minimum absolute average log2 ratio for region = 0.4 corresponding to a-fold change of 1.32; and, the ADM-2 algorithm was used for statistical analysis.