Variant filtering
Ten genes (MYOC, CYP1B1, LTBP2, PITX2, FOXC1, PAX6, TEK, ANGPT1, LMX1B, CPAMD8 ) known to cause various forms of early-onset glaucoma were initially screened for pathogenic mutations. If no plausible causative variants were identified, the exome data was examined for novel pathogenic variants. Disease-causing mutations are expected to be rare, disrupt protein function, and have high estimates of pathogenicity. Variants were filtered using 1) minor allele frequency (MAF) from the Exome Aggregation Consortium (ExAC) database (MAF less than 0.001 were retained), 2) tools to score variants according to phylogenetic conservation (GERP) and its effect on protein structure (SIFT, PolyPhen, conserved predicted pathogenic variants were obtained), 3) ocular expression, and any known functional or human disease involvement (ocular expression was prioritized) and 4) examining segregation in all available family members. The family A and B pedigree structures suggested dominant inheritance; hence, heterozygous variants were prioritized. Sanger sequencing was used to investigate segregation of candidate variants in all available family members.