Validation of a Custom 30-Gene Probe-Capture Targeted Sequencing Panel to Detect Variants in FEVR-Linked Genes and the RS1 Gene
Document Type
Conference Proceeding
Publication Date
6-2025
Publication Title
Investigative Ophthalmology and Visual Science
Abstract
Purpose : To validate a novel custom 30-gene probe-capture panel that includes genes linked to FEVR, Norrie Disease, Persistent Fetal Vascular Syndrome, and XLRS (Retinoschisis). Samples previously sequenced with a custom Ampliseq 8-gene panel (NDP, FZD4, LRP5, TSPAN12, KIF11, CTNNB1, ZNF408, RS1) were reanalyzed against the 30-gene probe-capture workflow.
Methods : DNA sequencing was carried out with the Oakland University Institutional Review Board and Associated Retinal Consultants (ARC) IRB approval. Subjects were consented to the ARC Eye Biobank, and DNA-sequencing in the Eye Research Institute. Six genomic DNA samples that were previously sequenced with an 8-gene Ampliseq panel were resequenced using a custom 30-gene probe capture panel (Celemics). The 30-gene and 8-gene Illumina-compatible libraries were sequenced in-house on an Illumina iSeq-100 instrument. DNA samples were from different years and extracted with a manual or automated DNA extraction kit. Variant analysis employed the Ensembl Variant Effect Predictor, ClinVar, DbSNP, and GnomAD to categorize variant consequence and estimate variant frequency in the population.
Results : The probe-capture 30-gene panel successfully identified 100% of the variants previously detected by the Ampliseq 8-gene panel in the following genes: LRP5 (1 variant), ZNF408 (2 variants), RS1 (1 variant), and NDP (1 variant). In addition, the 30-gene probe capture approach uncovered several novel variants not identified in the previous sequencing, demonstrating an enhanced capacity to detect a broader range of potentially pathogenic variants. The panel also detected a previously known RS1 gene variant.
Conclusions : These test results demonstrate that the newer custom Celemics probe-capture 30-gene panel can replace the previous 8-gene panel used by our group for DNA sequencing of FEVR-related genes and the RS1 gene. The expanded 30-gene panel also includes several additional genes more recently linked to FEVR such as CTNND1, CTNNA1, and ECM1.
Volume
66
Issue
8
First Page
1837
Last Page
1837
Recommended Citation
Krikor S, Dailey WA, Pinnock C, Joczyk V, Drenser KA, Mitton KP. Validation of a custom 30-gene probe-capture targeted sequencing panel to detect variants in FEVR-linked genes and the RS1 gene. Invest Ophthal Vis Sci. 2025 Jun;66(8):1827.
Comments
ARVO (Association for Research in Vision and Ophthalmology) Annual Meeting, May 4-8, 2025, Salt Lake City, UT