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Vaksali, Estonia

Verma A.,Dr G Venkataswamy Eye Research Institute | Perumalsamy V.,Pediatric Eye Clinic | Shetty S.,Pediatric Eye Clinic | Kulm M.,Asper Biotech | Sundaresan P.,Dr G Venkataswamy Eye Research Institute
PLoS ONE | Year: 2013

Background:Leber congenital amaurosis (LCA) is the most severe form of inherited retinal visual impairment in children. So far, mutations in more than 20 genes have been known to cause LCA and among them, RPE65 is a suitable candidate for gene therapy. The mutational screenings of RPE65 and other LCA genes are requisite in support of emerging gene specific therapy for LCA. Therefore, we have carried out a comprehensive LCA genes screening using a combined approach of direct sequencing and DNA microarray based Asper chip analysis.Methodology/Principal Findings:Thirty clinically diagnosed index LCA cases from Southern India were screened for coding and flanking intronic regions of RPE65 through direct sequencing. Among thirty, 25 cases excluded from RPE65 mutations were subjected to Asper chip analysis, testing 784 known pathogenic variations in 15 major LCA genes. In RPE65 screening, four different pathogenic variations including two novel (c.361insT & c.939T>A) and two known (c.394G>A & c.361delT) mutations were identified in five index cases. In the chip analysis, seven known pathogenic mutations were identified in six index cases, involving genes GUCY2D, RPGRIP1, AIPL1, CRX and IQCB1. Overall, 11 out of 30 LCA cases (36.6%) revealed pathogenic variations with the involvement of RPE65 (16.6%), GUCY2D (10%), RPGRIP1 (3.3%), AIPL1 (3.3%) and CRX & IQCB1 (3.3%).Conclusions/Significance:Our study suggests that such combined screening approach is productive and cost-effective for mutation detection and can be applied in Indian LCA cohort for molecular diagnosis and genetic counselling. © 2013 Verma et al.

Pereiro I.,University of Vigo | Hoskins B.E.,University College London | Marshall J.D.,The Jackson Laboratory | Collin G.B.,The Jackson Laboratory | And 6 more authors.
European Journal of Human Genetics | Year: 2011

Bardet-Biedl syndrome (BBS; OMIM no. 209 900) and Alström syndrome (ALMS; OMIM no. 203 800) are rare, multisystem genetic disorders showing both a highly variable phenotype and considerable phenotypic overlap; they are included in the emerging group of diseases called ciliopathies. The genetic heterogeneity of BBS with 14 causal genes described to date, serves to further complicate mutational analysis. The development of the BBS-ALMS array which detects known mutations in these genes has allowed us to detect at least one mutation in 40.5% of BBS families and in 26.7% of ALMS families validating this as an efficient and cost-effective first pass screening modality. Furthermore, using this method, we found two BBS families segregating three BBS alleles further supporting oligogenicity or modifier roles for additional mutations. We did not observe more than two mutations in any ALMS family. © 2011 Macmillan Publishers Limited All rights reserved.

Teek R.,University of Tartu | Kruustuk K.,University of Tartu | Zordania R.,Tallinn Childrens Hospital | Joost K.,Tallinn Childrens Hospital | And 7 more authors.
International Journal of Pediatric Otorhinolaryngology | Year: 2010

Objective: The purpose of this study was to determine the prevalence of c.35delG and p.M34T mutations in the GJB2 gene among children with early onset hearing loss and within a general population of Estonia. Methods: Using an arrayed primer extension assay, we screened 233 probands with early childhood onset hearing loss for 107 different mutations in the GJB2 gene. We then looked for the two most common mutations, c.35delG and p.M34T, in a population of 998 consecutively born Estonian neonates to determine the frequency of these mutations in the general population. Results: In 115 (49%) of the patients with early onset hearing loss, we found a mutation in at least one allele of the GJB2 gene. Seventy-three (31%) were homozygous for the c.35delG mutation, seven (3%) were homozygous for the p.M34T mutation, and five (2%) had c35delG/p.M34T compound heterozygosity. Other six identified mutations in GJB2 gene occurred rarely. Among the 998 anonymous newborn samples, we detected 45 who were heterozygous for c.35delG, 2 individuals homozygous for c.35delG, and 58 who were heterozygous for p.M34T. Additionally, we detected two c.35delG/p.M34T compound heterozygotes. Conclusion: The most common GJB2 gene mutations in Estonian children with early onset hearing loss were c.35delG and p.M34T, with c.35delG accounting for 75% of GJB2 alleles. The carrier frequency for c.35delG and p.M34T in a general population of Estonia was 1 in 22 and 1 in 17, respectively, and was higher than in most other countries. © 2010 Elsevier Ireland Ltd.

Vilbaste M.,University of Tartu | Slavin G.,Asper Biotech | Saks O.,University of Tartu | Pihl V.,University of Tartu | Leito I.,University of Tartu
Measurement: Journal of the International Measurement Confederation | Year: 2010

The GUM modelling, its Bayesian modification and the Monte Carlo method (MCM) to estimate the uncertainty are compared in two practical measurement situations (finding reference value of relative humidity and a generic chemical instrumental analysis procedure). The results of the three approaches agree very well when there are no dominant input quantities with type A evaluated uncertainty estimated from small number of repeated measurements. In the opposite case the GUM gives underestimated expanded uncertainties (by up to 20-25%), compared to both other approaches. Analysis of the practical measurement situations reveals that even in the case of several dominating input quantities of similar uncertainty contributions, if one of them is distributed according to the t-distribution and has a low number (3-4) of degrees of freedom, the output quantity cannot be safely assumed Normally distributed and in such a case coverage factor 2 is not an equivalent to 95% coverage level. © 2009 Elsevier Ltd. All rights reserved.

Pergament E.,Northwestern Reproductive Genetics | Alamillo C.,Northwestern Reproductive Genetics | Sak K.,Asper Biotech | Fiddler M.,DePaul University
Prenatal Diagnosis | Year: 2011

The objective of this study was to assess the first formal approach for monitoring genetic/developmental syndromes associated with the presence of an increased nuchal translucency (NT) thickness (>3 mm) in the first trimester of pregnancy. +Methods: Multiple technologies-a DNA chip using the APEX technology, qPCR, microfluidic PCR, and sequencing-were applied to assay 310 mutations across five conditions-Noonan syndrome, congenital adrenal hyperplasia, spinal muscular atrophy (SMA), DiGeorge syndrome, and Smith-Lemli Opitz syndrome. +Results: We report the results of assessing the first 120 patients in which 8 cases of Noonan syndrome were detected as well as an unusually high rate of heterozygosity for SMA. +Conclusion: While testing for Noonan syndrome in association with increased NT appears warranted, the reported association of the remaining four genetic syndromes is likely to be weak and possibly insignificant. © 2011 John Wiley & Sons, Ltd.

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