Sequence analysis of CYP21A1P in a German population to aid in the molecular biological diagnosis of congenital adrenal hyperplasia.

BACKGROUND: The high homology between the CYP21A2 (cytochrome P450, family 21, subfamily A, polypeptide 2) and CYP21A1P (cytochrome P450, family 21, subfamily A, polypeptide 1 pseudogene) genes is the major obstacle to risk-free genetic diagnosis of congenital adrenal hyperplasia, especially regarding the quantification of gene dosage. Because of the lack of a comprehensive study providing useful information about the detailed genetic structure of CYP21A1P, we used a large data set to analyze and characterize this pseudogene. METHODS: We amplified and directly sequenced the CYP21A1P and CYP21A2 genes of 200 unrelated individuals. The resulting sequence data were aligned against the manually curated transcript ENST0000448314 from Havana/Vega matching to the genebuild ENSG00000198457; all differences were documented. Copy number was measured by multiplex ligation-dependent probe amplification when necessary. RESULTS: We found that 40 potentially variable positions in CYP21A2 were conserved in CYP21A1P in all study participants. In addition, we detected 14 CYP21A1P variants that were not previously reported in either CYP21A2 or CYP21A1P. Unlike CYP21A2, CYP21A1P possessed certain mutation haplotypes. CONCLUSIONS: The genetic structure of CYP21A1P and the potential risks of false conclusions it may introduce are essential considerations in designing a PCR-based diagnosis procedure for congenital adrenal hyperplasia.