Detection of steroid 21-hydroxylase alleles using gene-specific PCR and a multiplexed ligation detection reaction
- Cornell Univ. Medical College, New York, NY (United States); and others
Steroid 21-hydroxylase deficiency is the most common cause of congenital adrenal hyperplasia, an inherited inability to synthesize cortisol that occurs in 1 in 10,000-15,000 births. Affected females are born with ambiguous genitalia, a condition that can be ameliorated by administering dexamethasone to the mother for most of gestation. Prenatal diagnosis is required for accurate treatment of affected females as well as for genetic counseling purposes. Approximately 95% of mutations causing this disorder result from recombinations between the gene encoding the 21-hydroxylase enzyme (CYP21) and a linked, highly homologous pseudogene (CYP21P). Approximately 20% of these mutations are gene deletions, and the remainder are gene conversions that transfer any of nine deleterious mutations from the CYP21P pseudogene to CYP21. We describe a methodology for genetic diagnosis of 21-hydroxylase deficiency that utilizes gene-specific PCR amplification in conjunction with thermostable DNA ligase to discriminate single nucleotide variations in a multiplexed ligation detection assay. The assay has been designed to be used with either fluorescent or radioactive detection of ligation products by electrophoresis on denaturing acrylamide gels and is readily adaptable for use in other disease systems. 30 refs., 5 figs.
- OSTI ID:
- 433296
- Journal Information:
- Genomics, Vol. 29, Issue 1; Other Information: PBD: 1 Sep 1995
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
BASIC STUDIES
HUMAN CHROMOSOME 6
GENETIC MAPPING
GENE MUTATIONS
DETECTION
HYDROXYLASES
GENE RECOMBINATION
METABOLIC DISEASES
DIAGNOSIS
HYDROCORTISONE
BIOSYNTHESIS
BIOASSAY
EVALUATION
SENSITIVITY
HISTOCOMPATIBILITY COMPLEX
GENES
POLYMERASE CHAIN REACTION
STEROIDS
CONGENITAL DISEASES
RADIOENZYMATIC ASSAY
EMBRYOS
DNA HYBRIDIZATION