The study demonstrates the successful application of exome sequencing to identify a gene for Kabuki syndrome, an autosomal dominant disorder characterized by distinctive facial features, cardiac anomalies, skeletal abnormalities, immunological defects, and mild to moderate mental retardation. Exomes of ten unrelated individuals with Kabuki syndrome were sequenced, and after filtering against SNP databases, no compelling candidate gene was identified in all affected individuals. However, less stringent filtering criteria identified multiple candidate genes, but genotypic and phenotypic stratification highlighted *MLL2*, a *Trithorax*-group histone methyltransferase, in which seven probands had novel nonsense or frameshift mutations. Follow-up Sanger sequencing confirmed *MLL2* mutations in two of the three remaining cases and in 26 of 43 additional cases. In families with available parental DNA, the mutation was confirmed to be *de novo* or transmitted. The results strongly suggest that mutations in *MLL2* are a major cause of Kabuki syndrome.The study demonstrates the successful application of exome sequencing to identify a gene for Kabuki syndrome, an autosomal dominant disorder characterized by distinctive facial features, cardiac anomalies, skeletal abnormalities, immunological defects, and mild to moderate mental retardation. Exomes of ten unrelated individuals with Kabuki syndrome were sequenced, and after filtering against SNP databases, no compelling candidate gene was identified in all affected individuals. However, less stringent filtering criteria identified multiple candidate genes, but genotypic and phenotypic stratification highlighted *MLL2*, a *Trithorax*-group histone methyltransferase, in which seven probands had novel nonsense or frameshift mutations. Follow-up Sanger sequencing confirmed *MLL2* mutations in two of the three remaining cases and in 26 of 43 additional cases. In families with available parental DNA, the mutation was confirmed to be *de novo* or transmitted. The results strongly suggest that mutations in *MLL2* are a major cause of Kabuki syndrome.