Molecular Genetic Study Of Congenital Idiopathic Nystagmus

ObjectiveTo study the clinical manifestations of three Chinese families(CZ,ZB and C) with X-linked C IN (congenital idiopathic nystagmus) and investigate genetic pathogenesis according to linkage analysis and mutation screening of candidate gene FRMD7.Methods1.Ophthalmologic examinations including best corrected visual acuity, refractive error, slit-lamp,fundus ophthalmoscopy, eye tracker system or VEP and ERG.2. Human genomic DNA was extracted from5-8ml peripheral blood leukocytes.Two microsatellites DXS1047and DXS1001at Xq26-q27were amplified by PCR reaction for linkage study. Two point analysis for these markers were calculated using the Milink program of Linkage package (version4.0).4. Direct genomic sequencingof FRMD7gene was conducted on all affected members in three families. Once the probable mutation was identified,100unaffected male individuals were screened to exclude the mutation as a polymorphism in a reference population.5.MLPA(Multiplex ligation-dependent probe amplification) analysis was carried out in family C using the supplied protocol from SALSA P269FRMD7-NYS1probe kit by MRC-Holland (Amsterdam, Holland).Results1.1Linkage study of family CZ showed that the maximum LOD score of1.91was yielded at the polymorphic marker DXS1001and a positive lod score of1.31at DXS1047(0=0). Sequencing of FRMD7gene showed a nucleotide change of c. 623A>G in the exon7of FRMD7gene in the patients, which predicted to result in an H208R amino acid change. This novel mutation was absent in100normal Han Chinese controls.1.2Linkage study of family ZB showed that the maximum LOD score of2.4was yielded at the polymorphic marker DXS1001and a positive lod score of1.5at DXS1047(0=0). Sequencing of FRMD7gene showed a nucleotide change of c.837G>C in the exon9of FRMD7gene in the patients, which predicted to result in an R279S amino acid change. This novel mutation was absent in100normal Han Chinese controls.1.3A significant lod score of2.4was yielded at the microsatellite marker DXS1047in family C. But no FRMD7gene mutations were detected by Sequencing and MLPA in the patients.Conclusion1. Three families are all X-linked CIN.2. We identified two novel mutation, c.837＞C (p. R279S) and c.623A>G (p. H208R), in two Han Chinese family with CIN. These mutations expand the mutation spectrum of FRMD7and help to further study molecular pathogenesis of FRMD7gene.3. The candidate region of disease gene of the family C was located on Xq26-Xq27. No mutations were detected by either direct sequencing or MLPA showed that the techniques of screening mutations in our experiments could be limited in detecting the unknown regions, for example, the promoter or other regulatory regions including those within introns, in the FRMD gene.