title: Novel method to characterize CYP21A2 in Florida patients with congenital adrenal hyperplasia and commercially available cell lines.
abstract: Congenital adrenal hyperplasia (CAH) is an autosomal recessive disorder and affects approximately 1 in 15,000 births in the United States. CAH is one of the disorders included on the Newborn Screening (NBS) Recommended Uniform Screening Panel. The commonly used immunological NBS test is associated with a high false positive rate and there is interest in developing second-tier assays to increase screening specificity. Approximately 90% of the classic forms of CAH, salt-wasting and simple virilizing, are due to mutations in the CYP21A2 gene. These include single nucleotide changes, insertions, deletions, as well as chimeric genes involving CYP21A2 and its highly homologous pseudogene CYP21A1P. A novel loci-specific PCR approach was developed to individually amplify the CYP21A2 gene, the nearby CYP21A1P pseudogene, as well as any 30 kb deletion and gene conversion mutations, if present, as single separate amplicons. Using commercially available CAH positive specimens and 14 families with an affected CAH proband, the single long-range amplicon approach demonstrated higher specificity as compared to previously published methods.
authors.affiliation: U.S. Centers for Disease Control and Prevention, National Center for Environmental Health, Division of Laboratory Sciences, Newborn Screening and Molecular Biology Branch, Atlanta, GA, United States.
authors.forename: Christopher N
authors.initials: CN
authors.lastname: Greene
authors.affiliation: U.S. Centers for Disease Control and Prevention, National Center for Environmental Health, Division of Laboratory Sciences, Newborn Screening and Molecular Biology Branch, Atlanta, GA, United States.
authors.forename: Suzanne K
authors.initials: SK
authors.lastname: Cordovado
authors.affiliation: U.S. Centers for Disease Control and Prevention, National Center for Environmental Health, Division of Laboratory Sciences, Newborn Screening and Molecular Biology Branch, Atlanta, GA, United States.
authors.forename: Daniel P
authors.initials: DP
authors.lastname: Turner
authors.affiliation: U.S. Centers for Disease Control and Prevention, National Center for Environmental Health, Division of Laboratory Sciences, Newborn Screening and Molecular Biology Branch, Atlanta, GA, United States.
authors.forename: Lisa M
authors.initials: LM
authors.lastname: Keong
authors.affiliation: University of South Florida, Department of Pediatrics, Tampa, FL, United States; All Children's Hospital, St. Petersburg, FL, United States.
authors.forename: Dorothy
authors.initials: D
authors.lastname: Shulman
authors.affiliation: U.S. Centers for Disease Control and Prevention, National Center for Environmental Health, Division of Laboratory Sciences, Newborn Screening and Molecular Biology Branch, Atlanta, GA, United States.
authors.forename: Patricia W
authors.initials: PW
authors.lastname: Mueller
keywords: CYP21A2; Congenital adrenal hyperplasia; DNA sequence; Mutation analysis