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SNP Genotyping (snp + genotyping)
Selected AbstractsNovel high-throughput SNP genotyping cosegregation analysis for genetic diagnosis of autosomal recessive retinitis pigmentosa and Leber congenital amaurosis,HUMAN MUTATION, Issue 5 2007Esther Pomares Abstract Retinitis pigmentosa (RP), the major cause of blindness in adults, is an extremely heterogeneous monogenic disorder. More than 32 causative genes have been identified, 18 of which are involved in autosomal recessive RP (arRP); however, more than 50% of the cases remain unassigned. There are no major causative genes identified for arRP nor any prevalent mutations, which make mutational screening of the already reported RP genes extremely time consuming and costly. Nonetheless, this step is unavoidable for genetic diagnosis of patients and potential carriers, and it is a prerequisite before approaching the identification of new RP genes and loci. We have designed an innovative high-throughput time- and cost-effective strategy for cosegregation analysis of 22 genes of arRP and Leber congenital amaurosis (LCA; an autosomal recessive retinal dystrophy that shares some of the RP genes and traits) by SNP genotyping. This novel indirect method has been validated in a panel of 54 consanguineous and nonconsanguineous arRP families. In a single and fast genotyping step: 1) we discarded all the 22 candidate genes in 13% of the pedigrees, highlighting the families of choice to search for novel arRP genes/loci; 2) we excluded an average of 18,19 genes per family, thus diminishing the number of genes to screen for pathogenic mutations; and 3) we identified CERKL as the causative RP gene in a family in which this candidate had been previously discarded by microsatellite cosegregation analysis. This type of approach can also be applied to other nonretinal diseases with high genetic heterogeneity, such as hereditary deafness or Parkinson disease. Hum Mutat 28(5), 511,516, 2007. © 2007 Wiley-Liss, Inc. [source] Novel pharmacogenetic markers for treatment outcome in azathioprine-treated inflammatory bowel diseaseALIMENTARY PHARMACOLOGY & THERAPEUTICS, Issue 4 2009M. A. SMITH Summary Background, Azathioprine (AZA) pharmacogenetics are complex and much studied. Genetic polymorphism in TPMT is known to influence treatment outcome. Xanthine oxidase/dehydrogenase (XDH) and aldehyde oxidase (AO) compete with TPMT to inactivate AZA. Aim, To assess whether genetic polymorphism in AOX1, XDH and MOCOS (the product of which activates the essential cofactor for AO and XDH) is associated with AZA treatment outcome in IBD. Methods, Real-time PCR was conducted for a panel of single nucleotide polymorphism (SNPs) in AOX1, XDH and MOCOS using TaqMan SNP genotyping assays in a prospective cohort of 192 patients receiving AZA for IBD. Results, Single nucleotide polymorphism AOX1 c.3404A > G (Asn1135Ser, rs55754655) predicted lack of AZA response (P = 0.035, OR 2.54, 95%CI 1.06,6.13) and when combined with TPMT activity, this information allowed stratification of a patient's chance of AZA response, ranging from 86% in patients where both markers were favourable to 33% where they were unfavourable (P < 0.0001). We also demonstrated a weak protective effect against adverse drug reactions (ADRs) from SNPs XDH c.837C > T (P = 0.048, OR 0.23, 95% CI 0.05,1.05) and MOCOS c.2107A > C, (P = 0.058 in recessive model, OR 0.64, 95%CI 0.36,1.15), which was stronger where they coincided (P = 0.019). Conclusion, These findings have important implications for clinical practice and our understanding of AZA metabolism. [source] A functional RANKL polymorphism associated with younger age at onset of rheumatoid arthritisARTHRITIS & RHEUMATISM, Issue 10 2010Wenfeng Tan Objective We previously observed the association of the co-occurrence of the HLA,DRB1 shared epitope (SE) and RANKL single-nucleotide polymorphisms (SNPs) with younger age at the onset of rheumatoid arthritis (RA) in 182 rheumatoid factor (RF),positive European American patients with early-onset RA. The aim of this study was to fine-map the 48-kb RANKL region in the extended cohort of 210 European American RF-positive patients with early RA, to seek replication of RA-associated SNPs in additional RA cohorts of 501 European Americans and 298 African Americans, and to explore the functional consequences of RA-associated SNPs. Methods SNP genotyping was conducted using pyrosequencing or TaqMan polymerase chain reaction (PCR) assays. Associations of rs7984870 with RANKL expression in plasma, peripheral blood mononuclear cells, and isolated T cells were quantified using enzyme-linked immunosorbent assay and reverse transcription,PCR. Site-directed mutagenesis of rs7984870 within the 2-kb RANKL promoter was performed to drive the luciferase reporter gene in osteoblast and stromal cell lines. Interaction of DNA and protein was determined by electrophoretic mobility shift assay. Results A single promoter SNP, rs7984870, was consistently significantly associated with earlier age at the onset of RA in 3 independent seropositive (RF or anti,cyclic citrullinated peptide antibody) RA cohorts but not in seronegative RA patients. The C risk allele of rs7984870 conferred 2-fold higher plasma RANKL levels in RF-positive patients with RA, significantly elevated RANKL messenger RNA expression in activated normal T cells, and increased promoter activity after stimulation in vitro via differential binding to the transcription factor SOX5. Conclusion The RANKL promoter allele that increased transcription levels upon stimulation might promote interaction between activated T cells and dendritic cells, predisposing to a younger age at the onset of RA in seropositive European American and African American patients. [source] 2242: Update on ophthalmic molecular geneticsACTA OPHTHALMOLOGICA, Issue 2010E DE BAERE Purpose To provide an overview of the recent technological advances in human molecular genetics that can be applied in ophthalmic genetics. Methods Since the finalization of the Human Genome Project many novel genomic technologies emerged that led to significant advances in gene identification and genetic testing of hereditary eye disorders: (1) genomewide copy number screening (array CGH); (2) genomewide SNP genotyping; (3) next-generation sequencing. Results (1) Microarray comparative genomic hybridisation or array CGH allows genomewide discovery of submicroscopic deletions and duplications in a single experiment. This technique is applied in routine molecular cytogenetic testing. Using array CGH a causal genomic defect can be found in at least 10% of all cases with mental retardation and/or multiple congenital anomalies. In ophthalmic genetics array CGH is mainly useful in the context of developmental eye disorders, with chorioretinal coloboma and anterior segment dysgenesis as an example. (2) Genomewide chip-based SNP genotyping can be used for homozygosity mapping in inbred and outbred pedigrees. Recent successes in gene identification using this approach are illustrated. (3) Next-generation sequencing or NGS. The application of this technology in gene identification and genetic testing of genetically heterogeneous conditions (with LCA as a paradigm) is discussed. Conclusion The rapid progress of genomic technologies such as array CGH, SNP chip analysis and next-generation sequencing lead to a boost in gene identification and genetic testing of both developmental and retinal eye disease. [source] Association of the interleukin-23 receptor gene variant rs11209026 with Crohn's disease in German childrenACTA PAEDIATRICA, Issue 5 2010M Lacher Abstract Aim:, Genome-wide association studies have described variants within the interleukin-23 receptor (IL23R) locus to be associated with Crohn's disease (CD) and ulcerative colitis (UC). We investigated the association of rs11209026 (p.Arg381Gln) and rs7517847 (c.799-3588T>G) into German paediatric inflammatory bowel disease (IBD) patients and analysed IL23R transcriptional activity in colonic tissues. Methods:, The rs11209026 and rs7517847 nucleotide substitutions were determined in 353 German children with IBD (221 CD, 132 UC) and 253 controls using pre-designed TaqMan® SNP genotyping assays. In selected IBD patients and controls, IL23R mRNA expression was measured using real-time PCR. Results:, The prevalence of the rs11209026 A allele was lower in CD patients, but not in UC patients, when compared with controls (1.8% vs 7.1%, p < 0.01). The rs7517847 variant, in contrast, was associated neither with CD nor with UC. IL23R expression was variable in IBD patients compared with controls without significant overexpression or downregulation. Conclusion:, Our study provides additional support for the strong protection of the rs11209026 (p.Arg381Gln) variant against paediatric CD. IL23R was expressed in both CD and UC with a great variability. However, expression levels showed no significant association with the disease. [source] | ||||||||||