High frequency of CRB1 mutations as cause of Early-Onset Retinal Dystrophies in the Spanish population

Table 1 General description of patients and molecular tools included in this study

Families in study	APEX microarray			Indirect studies		SSCP^&	HRM			Sanger sequencing	Total families characterized
	CRB1 2 alleles	Second allele by whole CRB1 analysis *	Total characterized	Haplotypes	IBD mapping	CRB1 2 alleles	CRB1 2 alleles	CRB1 1 allele	Second allele by Sanger sequencing *	2 alleles	CRB1 mutations	Mutations in another gene ^#
114 LCA	8 /114	4 / 8 ^a	12 (10%)	0 / 20	1^b/43	1 / 3	1 / 32	1 / 32	0 / 1	1/51	16 (14%)	32 (28%)
290 EORP	6 /290	15 /25 ^a	21 (7%)	0 / 60	0 / 23	1 / 20	0 / 193	1 / 193	1 / 1 ^c	4^d/ 209	27 (9%)	59 (20%)
404 Total	14 / 404	19 /33	33 (8%)	0 / 80	1 / 66	2 / 23	1 / 225	2 / 225	1 / 2	5/ 260	43 (11%)	91 (22%)

Overview of the sequential steps performed during the mutational CRB1 analysis in a cohort of LCA and EORP Spanish patients was reflected. The number of cases characterized and studied is outlined for each molecular tool. APEX: Arrayed primer extension; IBD: Inherited-by-descent; SSCP: single-strand conformation polymorphism analysis; HRM: High resolution melting analysis. * A direct mutational scanning of CRB1 was performed using dHPLC, HRM or Sanger sequencing in patients with a first allele identified by APEX microarray. ^& SSCP findings were reported by Bernal S. et als, 2003. ^# Mutations in another gene were found by APEX microarray, whole-genome homozygosity maping, whole exome sequencing or targeted NGS (data not shown).
^a A second CRB1 allele was not found in 14 patients: 2 carried a known frameshift mutation and 12 carried a uncertain or very unlikely missense variant.
^b A 5 Mb-homozygous region involving CRB1 was identified in an endogamic family and a homozygous known mutation was further found by Sanger sequencing. This mutation represents a false negative of the previously LCA chip analysis.
^c A second allele (c.1702C>T) was further found by Sanger sequencing, representing a false negative of the HRM analysis.
^d Two heterozygous transitions (c.2291 G>A and c.4168C>T) were found in one patient that previously showed normal melting curves in the HRM analysis thus, representing false negatives.

ISSN: 1750-1172