PCR-SSCP ANALYSIS OF TYPE-VII COLLAGEN GENE MUTATION IN DOMINANT DYSTROPHIC EPIDERMOLYSIS-BULLOSA

We performed SSCP (single-strand conformation polymorphism) and automated DNA sequencing to detect mutations of the Type VII collagen gene in two Japanese families suffering from dominant dystrophic epidermolysis bullosa (DDEB). Initially, using PCR (polymerase chain reaction), we amplified the NC1 domain of type VII collagen in genomic DNA obtained from either unaffected individuals or DDEB patients. SSCP analyses thereafter showed that int he DDEB patients a characteristic slower migrating band was present in addition to several bands identical to those found in unaffected individuals. Subsequent DNA sequencing established the presence of A-->G substitution (position 375) at one of the two alleles in all the DDEB patients from the two Japanese families. The above-described approaches are simple, quick, and require no radioactive materials, and therefore may be useful for screening gene mutations in congenital diseases. It could also be used to detect DDEB for example by prenatal diagnosis.