Development of a genotyping microarray for Usher syndrome.
BACKGROUND
Usher syndrome, a combination of retinitis pigmentosa (RP) and sensorineural hearing loss with or without vestibular dysfunction, displays a high degree of clinical and genetic heterogeneity. Three clinical subtypes can be distinguished, based on the age of onset and severity of the hearing impairment, and the presence or absence of vestibular abnormalities. Thus far, eight genes have been implicated in the syndrome, together comprising 347 protein-coding exons.


METHODS
To improve DNA diagnostics for patients with Usher syndrome, we developed a genotyping microarray based on the arrayed primer extension (APEX) method. Allele-specific oligonucleotides corresponding to all 298 Usher syndrome-associated sequence variants known to date, 76 of which are novel, were arrayed.


RESULTS
Approximately half of these variants were validated using original patient DNAs, which yielded an accuracy of >98%. The efficiency of the Usher genotyping microarray was tested using DNAs from 370 unrelated European and American patients with Usher syndrome. Sequence variants were identified in 64/140 (46%) patients with Usher syndrome type I, 45/189 (24%) patients with Usher syndrome type II, 6/21 (29%) patients with Usher syndrome type III and 6/20 (30%) patients with atypical Usher syndrome. The chip also identified two novel sequence variants, c.400C>T (p.R134X) in PCDH15 and c.1606T>C (p.C536S) in USH2A.


CONCLUSIONS
The Usher genotyping microarray is a versatile and affordable screening tool for Usher syndrome. Its efficiency will improve with the addition of novel sequence variants with minimal extra costs, making it a very useful first-pass screening tool.
Cremers
Frans P M
FP
Department of Human Genetics, and Nijmegen Centre for Molecular Life Sciences, Radboud University Nijmegen Medical Centre, Nijmegen, The Netherlands. F.Cremers@antrg.umcn.nl
Kimberling
William J
WJ
Külm
Maigi
M
de Brouwer
Arjan P
AP
van Wijk
Erwin
E
te Brinke
Heleen
H
Cremers
Cor W R J
CW
Hoefsloot
Lies H
LH
Banfi
Sandro
S
Simonelli
Francesca
F
Fleischhauer
Johannes C
JC
Berger
Wolfgang
W
Kelley
Phil M
PM
Haralambous
Elene
E
Bitner-Glindzicz
Maria
M
Webster
Andrew R
AR
Saihan
Zubin
Z
De Baere
Elfride
E
Leroy
Bart P
BP
Silvestri
Giuliana
G
McKay
Gareth J
GJ
Koenekoop
Robert K
RK
Millan
Jose M
JM
Rosenberg
Thomas
T
Joensuu
Tarja
T
Sankila
Eeva-Marja
EM
Weil
Dominique
D
Weston
Mike D
MD
Wissinger
Bernd
B
Kremer
Hannie
H
D004247:DNA / Q000235:genetics; D017931:DNA Primers; D005060:Europe; D014644:Genetic Variation; D005838:Genotype; D006801:Humans; D020411:Oligonucleotide Array Sequence Analysis; D052245:Usher Syndromes / Q000235:genetics*
D017931:DNA Primers; D004247:DNA