CNV Analysis
CNVs were detected with our analytical pipeline of Illumina 1M arrays (v.1 and v.3)6,30 and analyzed for case-control differences in burden with PLINK v.1.0730, R stats, and custom scripts. The p values associated with odds ratios (ORs) were calculated with Fisher’s exact test. Rare de novo CNVs, clinically relevant CNVs, and other selected rare CNVs were validated by at least one method (quantitative PCR, MLPA, and/or long-range PCR). Table S4 shows all validated de novo CNVs. A list of CNV calls passing QC in affected subjects, including all experimentally validated CNVs, is available in Tables S17A, S17B, and S17C.
Secondary analyses included comparisons of CNV number, length, and intersected gene number between our 102 de novo CNVs identified in affected subjects and the 76 de novo CNVs in control subjects of two published data sets: (1) 17 de novo CNVs identified in 15 unaffected siblings from 872 families with a single ASD-affected offspring and an unaffected sibling from the Simons Simplex Collection4 and (2) 59 de novo CNVs detected in 57 out of 2,623 Icelandic control trios.31
The clinical relevance of CNVs was interpreted according to the American College of Medical Genetics guidelines32 irrespective of the subjects’ affected status, and CNVs were classified as pathogenic, uncertain, or benign. Pathogenic CNVs are documented as clinically significant in multiple peer-reviewed publications and databases (e.g., OMIM and GeneReviews), even if the penetrance and the expressivity might be variable.