title: Screening for CHARGE syndrome mutations in the CHD7 gene using denaturing high-performance liquid chromatography.
abstract: Mutations in the CHD7 (chromodomain helicase DNA binding protein 7) gene cause CHARGE syndrome. At present, however, genetic testing of the CHD7 gene is not commonly applied in clinical settings because the currently available assays are technically and financially demanding, mainly because of the size of the gene. In the present study, we optimized the highly sensitive and specific mutation scanning method automated denaturing high-performance liquid chromatography (DHPLC) to analyze the entire coding region of CHD7. The coding region was amplified by 39 primer pairs, all of which have the same cycling conditions, aliquoted on a 96-well format polymerase chain reaction (PCR) plate. In this manner, all of the exons were amplified simultaneously using a single block in a thermal cycler. We then wrote a computer script to analyze each segment of the CHD7 gene by DHPLC in a serial manner using conditions that were optimized for each amplicon. The implementation of this screening method for CHD7 will help medical geneticists confirm their clinical impressions and provide accurate genetic counseling to the patients with CHARGE syndrome and their families.
authors.affiliation: Department of Pediatrics, Keio University School of Medicine, Tokyo, Japan.
authors.forename: Michihiko
authors.initials: M
authors.lastname: Aramaki
authors.forename: Toru
authors.initials: T
authors.lastname: Udaka
authors.forename: Chiharu
authors.initials: C
authors.lastname: Torii
authors.forename: Hazuki
authors.initials: H
authors.lastname: Samejima
authors.forename: Rika
authors.initials: R
authors.lastname: Kosaki
authors.forename: Takao
authors.initials: T
authors.lastname: Takahashi
authors.forename: Kenjiro
authors.initials: K
authors.lastname: Kosaki
mesh_terms: D000015:Abnormalities, Multiple / Q000235:genetics*; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D004265:DNA Helicases / Q000235:genetics*; D004252:DNA Mutational Analysis; D004268:DNA-Binding Proteins / Q000235:genetics*; D004427:Ear Diseases / Q000235:genetics; D005128:Eye Diseases / Q000235:genetics; D005820:Genetic Testing / Q000379:methods*; D006330:Heart Defects, Congenital / Q000235:genetics; D006801:Humans; D009668:Nose Diseases / Q000235:genetics; D009691:Nucleic Acid Denaturation; D016133:Polymerase Chain Reaction; D013577:Syndrome
chemical_list: D004268:DNA-Binding Proteins; D004265:DNA Helicases; C491434:CHD7 protein, human