Development of a Genotyping Microarray for Usher Syndrome

Abstract

Abstract: : Purpose: Patients with Usher syndrome are classified in three clinical subtypes and 8 causative genes have been identified. We set out to develop a robust and cost–effective mutation screening platform for Usher syndrome based on allele–specific primer extension. Methods: All (296) known USH variants residing in the CDH23, MYO7A, PCDH15, USH1C, USH1G, USH2A, USH3A, and VLGR1 genes were collected. Allele–specific and exon–flanking oligonucleotides were designed. For the validation and evaluation of the chip, DNAs from 150 and 375 patients respectively were employed. 175 of the latter patients were novel; 200 patients were selected from an USH1 patient cohort in which no MYO7A mutations were found and an USH2 patient cohort in which the frequent USH2A p.E767fs (c.2299delG) variant was not found. Results: After troubleshooting, the USH chip is estimated to be 95% accurate for the known sequence variants. The analysis of novel Usher syndrome DNA samples revealed putative pathologic variants in ∼50% of USH1 alleles and in ∼20% of USH2 alleles. In addition, sequence variants were identified in a large proportion of patients with atypical Usher syndrome. Sequence variants in USH1 genes were identified in USH2 patients and vice versa, the significance of which remains to be established. Conclusions: The novel Usher syndrome mutation chip for the first time enables a comprehensive and affordable analysis of all known sequence variants. For Usher syndrome type 1 patients and atypical cases, the efficiency of the chip is equal or superior to currently employed mutation scanning techniques. For Usher syndrome type 2, additional variants need to be identified and added to the chip to improve its efficiency.

Keywords: genetics • retina • photoreceptors