title: Rapid one-step carrier detection assay of mucolipidosis IV mutations in the Ashkenazi Jewish population.
abstract: Two mutations in the MCOLN1 mucolipidosis IV (ML IV) gene represent approximately 95% of the mutations in Ashkenazi-Jewish patients with ML IV. The mutations, a splice site mutation (IVS3-2A>G) and an approximately 6.4-kb deletion (511del6434), account for 72% and 23% of ML IV alleles in this population, respectively. An automated high-throughput assay was developed using the 5'-nuclease (TaqMan) method for the simultaneous detection of both mutations in a single reaction. Three fluorescent probes specifically detected wild-type, IVS3-2A>G, and 511del6434 alleles in each reaction real-time. Data collected were automatically analyzed, and genotype results were uploaded into a laboratory information management system. The assay was validated using genomic controls, demonstrating high robustness and accuracy. Carrier screening of 10,527 samples revealed 77 heterozygote carriers of IVS3-2A>G, 25 heterozygote carriers of 511del6434, and two compound heterozygote of both mutant alleles. The frequency of mutated alleles was 0.73% for IVS3-2A>G and 0.24% for 511del6434. The combined carrier frequency was 1:103 with predicted disease incidence of 1:42,436 individuals in this population, slightly lower than previously described frequencies. This automated high-throughput assay is labor saving, because two mutations can be detected in a single reaction. The method has potential for use in other assays requiring simultaneous detection of two mutations.
authors.affiliation: Department of Molecular Genetics, Quest Diagnostics Incorporated, Nichols Institute, San Juan Capistrano, California 92690, USA. feras.x.hantash@questdiagnostics.com
authors.forename: Feras M
authors.initials: FM
authors.lastname: Hantash
authors.forename: Susan C
authors.initials: SC
authors.lastname: Olson
authors.forename: Ben
authors.initials: B
authors.lastname: Anderson
authors.forename: Arlene
authors.initials: A
authors.lastname: Buller
authors.forename: Rebecca
authors.initials: R
authors.lastname: Chen
authors.forename: Beryl
authors.initials: B
authors.lastname: Crossly
authors.forename: Weimin
authors.initials: W
authors.lastname: Sun
authors.forename: Charles M
authors.initials: CM
authors.lastname: Strom
mesh_terms: D000483:Alleles; D004252:DNA Mutational Analysis / Q000379:methods*; D005820:Genetic Testing / Q000379:methods*; D006579:Heterozygote; D006801:Humans; D007585:Jews / Q000208:ethnology* / Q000235:genetics*; D009081:Mucolipidoses / Q000235:genetics*; D009154:Mutation / Q000235:genetics*; D013997:Time Factors