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Editor—Pseudoxanthoma elasticum (PXE, MIM 264800) is an inherited disorder of connective tissue in which the elastic fibres of the skin, eyes, and cardiovascular system slowly become calcified, causing a spectrum of disease involving these three organ systems, with highly variable phenotypic expression.1 2Mutations in the ABCC6 gene (previously known as MRP6), encoding a 1503 amino acid membrane transporter, have recently been identified by our group and others3-7 as the genetic defect responsible for PXE. We subsequently designed a strategy for a complete mutational analysis of the ABCC6 gene, in order to provide accurate molecular and prenatal diagnosis of PXE. During this mutational screening, we have found evidence for the existence of at least one pseudogene highly homologous to the 5′ end ofABCC6. Sequence variants in thisABCC6-like pseudogene could be mistaken for mutations in the ABCC6 gene and consequently lead to erroneous genotyping results in pedigrees affected with pseudoxanthoma elasticum.
Material and methods
Seven unrelated patients presenting with PXE were evaluated for mutational analysis of the ABCC6 gene. For each proband, diagnosis of PXE was consistent with previously reported consensus criteria,8 which include a positive von Kossa stain of a skin biopsy, indicating calcification of elastic fibres, in combination with specific cutaneous and ocular manifestations (angioid streaks).
Whole blood samples were obtained after participants had provided written consent using a form that was approved by the Institutional Review Board of our academic institution. High molecular weight DNA was isolated from peripheral blood leucocytes, using a standard salting out procedure. Primers for amplification of theABCC6 gene were designed from the published sequence of human chromosome 16 bacterial artificial chromosome (BAC) clone A-962B4 (GenBank accession number U91318). PCR amplifications ofABCC6 exon 2 and exon 9 were done in 20 μl volumes with 100 nmol/l …