title: CYP21A2 mutation analysis in Korean patients with congenital adrenal hyperplasia using complementary methods: sequencing after long-range PCR and restriction fragment length polymorphism analysis with multiple ligation-dependent probe amplification assay.
abstract: CYP21A2 mutation analysis of congenital adrenal hyperplasia (CAH) is challenging because of the genomic presence of a homologous CYP21A2 pseudogene and the significant incidence of pseudogene conversion and large deletions. The objective of this study was to accurately analyze the CYP21A2 genotype in Korean CAH patients using a combination of complementary methods. Long-range PCR and restriction fragment length polymorphism analyses were performed to confirm valid amplification of CYP21A2 and to detect large gene conversions and deletions before direct sequencing. Multiple ligation-dependent probe amplification (MLPA) analysis was conducted concurrently in 14 CAH-suspected patients and six family members of three patients. We identified 27 CYP21A2 mutant alleles in 14 CAH-suspected patients. The c.293-13A>G (or c.293-13C>G) was the most common mutation, and p.Ile173Asn was the second, identified in 25% and 17.9% of alleles, respectively. A novel frame-shift mutation of c.492delA (p.Glu 164Aspfs*24) was detected. Large deletions were detected by MLPA in 10.7% of the alleles. Mutation studies of the six familial members for three of the patients aided in the identification of haplotypes. In summary, we successfully identified CYP21A2 mutations using both long-range PCR and sequencing and dosage analyses. Our data correspond relatively well with the previously reported mutation spectrum analysis.
authors.affiliation: Department of Laboratory Medicine and Genetics, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Korea.
authors.forename: Geehay
authors.initials: G
authors.lastname: Hong
authors.affiliation: Department of Laboratory Medicine and Genetics, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Korea. nayadoo@hanmail.net.
authors.forename: Hyung Doo
authors.initials: HD
authors.lastname: Park
authors.affiliation: Department of Laboratory Medicine and Genetics, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Korea.
authors.forename: Rihwa
authors.initials: R
authors.lastname: Choi
authors.affiliation: Department of Pediatrics, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Korea.
authors.forename: Dong Kyu
authors.initials: DK
authors.lastname: Jin
authors.affiliation: Department of Medicine, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Korea.
authors.forename: Jae Hyeon
authors.initials: JH
authors.lastname: Kim
authors.affiliation: Department of Laboratory Medicine and Genetics, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Korea.
authors.forename: Chang Seok
authors.initials: CS
authors.lastname: Ki
authors.affiliation: Department of Laboratory Medicine and Genetics, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Korea.
authors.forename: Soo Youn
authors.initials: SY
authors.lastname: Lee
authors.affiliation: Department of Laboratory Medicine, Seoul National University Bundang Hospital, Seoul National University College of Medicine, Seongnam, Korea.
authors.forename: Junghan
authors.initials: J
authors.lastname: Song
authors.affiliation: Department of Laboratory Medicine and Genetics, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Korea.
authors.forename: Jong Won
authors.initials: JW
authors.lastname: Kim
mesh_terms: D000293:Adolescent; D000312:Adrenal Hyperplasia, Congenital / Q000175:diagnosis / Q000235:genetics*; D000328:Adult; D002648:Child; D004252:DNA Mutational Analysis; D005260:Female; D006801:Humans; D007223:Infant; D007231:Infant, Newborn; D022521:Ligase Chain Reaction; D008297:Male; D008875:Middle Aged; D016133:Polymerase Chain Reaction; D012150:Polymorphism, Restriction Fragment Length; D056910:Republic of Korea; D013255:Steroid 21-Hydroxylase / Q000235:genetics*; D055815:Young Adult
chemical_list: C531510:CYP21A2 protein, human; D013255:Steroid 21-Hydroxylase
keywords: CYP21A2; Korea; Multiple ligation-dependent probe amplification; Pseudogene; Restriction fragment length polymorphism