June 2013
Volume 54, Issue 15
ARVO Annual Meeting Abstract  |   June 2013
RNA-Seq Approach for the Refinement of a Modifier Locus in a Canine Model of Cone-Rod Dystrophy
Author Affiliations & Notes
  • Keiko Miyadera
    School of Vet Medicine, Univ of Pennsylvania, Philadelphia, PA
  • Matthew Brooks
    National Eye Institute, NIH, Bethesda, MD
  • Gautami Das
    School of Vet Medicine, Univ of Pennsylvania, Philadelphia, PA
  • Anand Swaroop
    National Eye Institute, NIH, Bethesda, MD
  • Gustavo Aguirre
    School of Vet Medicine, Univ of Pennsylvania, Philadelphia, PA
  • Footnotes
    Commercial Relationships Keiko Miyadera, None; Matthew Brooks, None; Gautami Das, None; Anand Swaroop, None; Gustavo Aguirre, None
  • Footnotes
    Support None
Investigative Ophthalmology & Visual Science June 2013, Vol.54, 3353. doi:
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      Keiko Miyadera, Matthew Brooks, Gautami Das, Anand Swaroop, Gustavo Aguirre; RNA-Seq Approach for the Refinement of a Modifier Locus in a Canine Model of Cone-Rod Dystrophy. Invest. Ophthalmol. Vis. Sci. 2013;54(15):3353.

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      © ARVO (1962-2015); The Authors (2016-present)

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Purpose: We have previously established a canine model of oligogenic cone-rod dystrophy (cord1). The primary locus represented by a homozygous insertion in RPGRIP1 (RPGRIP1ins/ins) was mapped in an early-onset cord1 research colony. A proportion of RPGRIP1ins/ins pet dogs, however, showed a slower disease progression with much later or no clinical onset. Subsequent GWAS in these RPGRIP1ins/ins dogs mapped a 1.5Mb homozygous modifier interval associated with early onset. As Sanger sequencing of positional candidate genes failed to identify a mutation, we have used now a transcriptomics approach to further refine the modifier locus.

Methods: RNA-seq was performed using retinal tissues from seven dogs in a newly established colony where the primary and the modifier loci segregated independently. High-throughput sequencing reads of retinal cDNA librarys were assembled to the canine reference. Differential expression of annotated genes/transcripts and unresolved transcribed sequences was analyzed. Based on sequence variants, haplotypes were constructed across the modifier interval.

Results: Of 12 annotated genes in the modifier interval, GUCY1A3 and GUCY1B3 were highly expressed in the retina; less abundantly expressed were MAP9 and CTSO. Other known genes showed trace or no retinal expression except for LRAT affected by RPE contamination. Homozygosity for the early-onset haplotype (modE/E) was confirmed in one RPGRIP1+/ins dog. One wildtype dog was homozygous only for a telomeric portion of the E haplotype (modE/TE). There were no double-homozygotes (RPGRIP1ins/insmodE/E) in the study samples, and early-onset cord1 was not observed clinically. There was no significant difference in the expression of annotated genes/transcripts along the modifier interval associated with the E haplotype. Among the unresolved regions, differential splicing pattern was found flanking the 3’-UTR of MAP9.

Conclusions: RNA-seq using canine retinal samples could be applied to assess differential expression of annotated and unannotated transcripts across the target interval. Sequence variants enabled constructing of haplotypes to determine identity by descent and chromosomal break points. There were no critical changes in sequences or expression levels corresponding to the E haplotype. The role of the MAP9 transcript variant in the interaction with the primary cord1 locus is being explored.

Keywords: 696 retinal degenerations: hereditary • 536 gene modifiers • 539 genetics  

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