We amplified coding regions in SYNGAP1 (National Center for Biotechnology Information build number, 36.1) and their intronic flanking regions using a polymerase-chain-reaction (PCR) assay of genomic DNA and then sequenced the resulting products. PCR primers targeting the 19 exons of SYNGAP1 were designed with the use of Exon-Primer from the University of California, Santa Cruz, Genome Browser (Table 1 in the Supplementary Appendix). PCR assays were performed in 384 well plates with the use of 5 ng of genomic DNA, according to standard procedures. PCR products were sequenced at the McGill University and Genome Quebec Innovation Centre in Montreal (www.genomequebecplatforms.com/mcgill) on a 3730XL DNA analyzer. In each case, unique mutations were confirmed by reamplification of the fragment and resequencing of the proband and both parents with the use of reverse and forward primers. PolyPhred (version 5.04), PolyScan (version 3.0), and Mutation Surveyor (version 3.10) were used for mutation-detection analyses.
