Comprehensive Chromosomal Copy Number Analysis

Analysis of CNAs was performed according to our previous reports [21 (link), 22 (link)]. Extracted DNA was adjusted to a concentration of 50 ng/μL. All 85 paired samples were assayed with use of the Infinium HumanCytoSNP-12 v2.1 BeadChip (Illumina, San Diego, CA, USA), which contains 299,140 SNP loci, according to the Illumina Infinium HD assay protocol. BeadChips were scanned by iScan (Illumina) and analysed by GenomeStudio (version 2011.1; Illumina). The log R ratio (LRR) and B allele frequency (BAF) for each sample were exported from normalized Illumina data with use of GenomeStudio. Data were analysed with use of KaryoStudio 1.4.3 (CNV Plugin version 3.0.7.0; Illumina) with default parameters. Chromosomal CNAs were classified by copy number variation partition algorithms: LRR = 0 indicated a normal diploid region, LRR > 0 indicated a copy number gain and LRR < 0 indicated a copy number LOH. BAFs ranged from 0 to 1; homozygous SNPs had BAFs near 0 (A allele) or 1 (B allele), and heterozygous diploid region SNPs had BAFs near 0.5 (AB genotype). Additionally, LRR and BAF data were used to identify regions of hemizygous and copy-neutral LOH.