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Sara J. Bowne, Lori S. Sullivan, Jennifer D. Churchill, Susan H. Blanton, Dianna K. Wheaton, David G. Birch, Kari E. Branham, John R. Heckenlively, Eric A. Pierce, Stephen P. Daiger; Genome-Wide Linkage Analysis For Gene Discovery In Autosomal Dominant Retinitis Pigmentosa. Invest. Ophthalmol. Vis. Sci. 2012;53(14):4528.
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© ARVO (1962-2015); The Authors (2016-present)
To use linkage exclusion and genome-wide linkage mapping in large adRP families to identify genomic regions containing unidentified adRP genes and mutations. Exome sequencing has been performed on these families and is currently focused on the mapped linkage regions, thereby reducing the number of variants requiring subsequent analysis and increasing the speed of mutation identification.
Ten families were selected from our well characterized cohort of 244 adRP families based on family size and availability of DNA samples. Families studied had from six to thirteen affected members available for testing. Each of the families had been tested previously for mutations in the known adRP genes and no mutations were found. STR markers located in or near the known adRP genes were analyzed to further exclude occult mutations as causes of disease. Genome-wide linkage analysis was performed on six of the families with DNA available from nine or more affected members. DNA was typed with Affymetrix 6.0 SNP chips and analyzed with MERLIN. Additional STR markers were run to exclude or validate any regions identified by linkage exclusion or SNP analyses.
A maximum LOD score greater than 3.0 was obtained for five of the families tested. Linkage exclusion analyses identified two families, RFS132 and UTAD055 with linkage to regions harboring known adRP genes on chromosome 19q13. MLPA and sequencing analysis of the CRX and PRPF31 genes in RFS132 and UTAD055 failed to identify pathogenic mutations in these families. Linkage exclusion analysis in UTAD598 led to identification of a new adRP locus on chromosome 2q12-q31 which is close to, but distinct from, the RP33 gene, SNRNP200. This new adRP locus, RP52, is in a region 68mB in size. Testing excluded the known adRP loci in RFS016 and UTAD392. Genome-wide linkage analysis in UTAD569 and UTAD562 led to the identification of two novel adRP loci. The disease locus in UTAD569 mapped to chromosome 1p and was determined to be caused by a dominant acting Asp477Gly mutation in RPE65. The disease locus in UTAD562 maps to chromosome 20q13.33. Exome and dideoxy sequencing suggest that the known adRP gene found in this region, PRPF6, is not the cause of disease in this family.
Linkage analysis is an effective tool for localizing the disease-causing mutation in adRP. Our analysis indicates that it is likely that new adRP genes are present on chromosome 2q, 19q and 20q. Genome-wide linkage data is still being analyzed for the RFS403, UTAD388, and UTAD076 families with the expectation that they too will indentify new adRP loci. This new linkage data is being combined with exome and whole genome sequencing data to more efficiently identify the disease-causing mutations in these families.
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