SIMPLIFIED DETECTION OF A MUTATION CAUSING FAMILIAL HYPERCHOLESTEROLEMIA THROUGHOUT BRITAIN - EVIDENCE FOR AN ORIGIN IN A COMMON DISTANT ANCESTOR

Familial hypercholesterolaemia (FH) is an inherited autosomal codomina nt disorder caused by many different mutations in the Io low-density l ipoprotein receptor (LDLR) gene. The one described most frequently in patients with FH from England, arises from a G-->A transition al the f irst nucleotide of codon 80, resulting ill the substitution of lysine for glutamic acid at residue SO of the mature protein, FH E80K. We des cribe a simple method to detect this mutation in genomic DNA using the polymerase chain reaction (PCR). A 69 base pair (bp) fragment of exon 3 of tile LDLR gene is amplified using a mutagenic upstream PCR prime r. This substitutes a T For an A residue in the amplified product, 2 b p upstream from the mutant site, generating a restriction site for the endonuclease Taq Il in normal but not in mutant DNA. Following digest ion of amplified DNA with Taq I, normal but not mutant DNA is cut into two fragments of 29 and 40 bp, which are readily identified by polyac rylamide gel electrophoresis. Using this method, 410 patients with cli nically diagnosed FH, attending lipid clinics in Edinburgh (72), Newpo rt (158), Walsall (30) and Southampton (150), were screened for the mu tation. Five individuals tasted positive as heterozygotes, one from Ed inburgh three from Newport and one from Southampton. This finding was confirmed by DNA sequence analysis. We conclude that FH due to this mu tation occurs in individuals throughout Great Britain and that it can be detected accurately using this simple technique. DNA from these and other individuals previously identified to be heterozygous for FH E80 K, was then studied using PCR of highly informative microsatellite mar kers flanking the LDLR gene. Sixteen of 17 apparently unrelated indivi duals heterozygous for FH E80K also were heterozygous for an identical size (239 nucleotide) allele, of polymorphic microsatellite D19S394. located approximately 250 kb away from the LDLR gene. This supports th e hypothesis that FH E80K in these 16 individuals arose from a single ancestor less than 1000 years ago.