Discrimination between metachromatic leukodystrophy and pseudo-deficiency of arylsulfatase A by restriction digest of amplified gene fragments.
Mutations causing metachromatic leukodystrophy and pseudo-deficiency were detected in the arylsulfatase A gene by methods based on different wild-type and mutant restriction sites. After polymerase chain reaction amplification of fragments of the arylsulfatase A gene and digestion by the appropriate endonuclease, the mixtures were separated by polyacrylamide gel electrophoresis and visualized by ethidium bromide staining. The common splice mutation in intron 2 (459 + 1G-->A) causing, in homozygosity, late-infantile metachromatic leukodystrophy and the common missense mutation in exon 8 (P426L) causing, in homozygosity, adult or juvenile metachromatic leukodystrophy were found to abolish Bst NI and Aci I sites, respectively. The polyadenylation pseudo-deficiency mutation (1619A-->G) was found to create a Mae III restriction site. The N-glycosylation pseudo-deficiency mutation (N350S) does not produce or destroy any known restriction site, and in this case, introduction of a single nucleotide mismatch in one of the primers enabled the authors to create a Bfa I site in the mutant allele.
Ben-Yoseph
Y
Y
Department of Pediatrics, Wayne State University School of Medicine, Detroit, Michigan.
Mitchell
D A
DA
D000483:Alleles; D001483:Base Sequence; D002553:Cerebroside-Sulfatase / Q000172:deficiency* / Q000235:genetics; D017931:DNA Primers; D003937:Diagnosis, Differential; D005091:Exons; D006801:Humans; D007962:Leukocytes / Q000201:enzymology; D007966:Leukodystrophy, Metachromatic / Q000175:diagnosis* / Q000201:enzymology / Q000235:genetics; D008969:Molecular Sequence Data; D017354:Point Mutation; D016133:Polymerase Chain Reaction; D015183:Restriction Mapping; D012867:Skin / Q000201:enzymology
D017931:DNA Primers; D002553:Cerebroside-Sulfatase