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Ophthalmic Genet. 2015;36(4):333-8. doi: 10.3109/13816810.2014.886269. Epub 2014 Feb 19.

Advantage of Whole Exome Sequencing over Allele-Specific and Targeted Segment Sequencing in Detection of Novel TULP1 Mutation in Leber Congenital Amaurosis.

Author information

1
a Center for Applied Genomics, The Children's Hospital of Philadelphia , Philadelphia , PA , USA .
2
b Eye and Developmental Genetics Research Group, Western Sydney Genetics Program, The Children's Hospital at Westmead , Sydney , NSW , Australia .
3
c Children's Medical Research Institute , Westmead , Sydney , NSW , Australia .
4
d College of Life Sciences, Sichuan University, Key Laboratory for Bio-resources and Eco-environment of Ministry of Education, Sichuan Key Laboratory of Molecular Biology and Biotechnology , Chengdu , PR China .
5
e BGI-Shenzhen , Shenzhen , China .
6
f Department of Ophthalmology , The Children's Hospital at Westmead , Sydney , NSW , Australia .
7
g Division of Human Genetics , The Children's Hospital of Philadelphia , Philadelphia , PA , USA .
8
h Department of Pediatrics , The Perelman School of Medicine, University of Pennsylvania , Philadelphia , PA , USA .
9
i Department of Biology , University of Copenhagen , Copenhagen , Denmark .
10
j King Abdulaziz University , Jeddah , Saudi Arabia .
11
k Discipline of Ophthalmology & Save Sight Institute, University of Sydney , Sydney , Australia , and.
12
l Disciplines of Paediatrics and Child Health & Genetic Medicine, University of Sydney , Sydney , NSW , Australia.

Abstract

BACKGROUND:

Leber congenital amaurosis (LCA) is a severe form of retinal dystrophy with marked underlying genetic heterogeneity. Until recently, allele-specific assays and Sanger sequencing of targeted segments were the only available approaches for attempted genetic diagnosis in this condition. A broader next-generation sequencing (NGS) strategy, such as whole exome sequencing, provides an improved molecular genetic diagnostic capacity for patients with these conditions.

MATERIALS AND METHODS:

In a child with LCA, an allele-specific assay analyzing 135 known LCA-causing variations, followed by targeted segment sequencing of 61 regions in 14 causative genes was performed. Subsequently, exome sequencing was undertaken in the proband, unaffected consanguineous parents and two unaffected siblings. Bioinformatic analysis used two independent pipelines, BWA-GATK and SOAP, followed by Annovar and SnpEff to annotate the variants.

RESULTS:

No disease-causing variants were found using the allele-specific or targeted segment Sanger sequencing assays. Analysis of variants in the exome sequence data revealed a novel homozygous nonsense mutation (c.1081C > T, p.Arg361*) in TULP1, a gene with roles in photoreceptor function where mutations were previously shown to cause LCA and retinitis pigmentosa. The identified homozygous variant was the top candidate using both bioinformatic pipelines.

CONCLUSIONS:

This study highlights the value of the broad sequencing strategy of exome sequencing for disease gene identification in LCA, over other existing methods. NGS is particularly beneficial in LCA where there are a large number of causative disease genes, few distinguishing clinical features for precise candidate disease gene selection, and few mutation hotspots in any of the known disease genes.

KEYWORDS:

Leber congenital amaurosis; TULP1; retinitis pigmentosa; whole exome sequencing

PMID:
24547928
DOI:
10.3109/13816810.2014.886269
[Indexed for MEDLINE]
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