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Title:High-resolution melting curve analysis for high-throughput genotyping of NOD2/CARD15 mutations and distribution of these mutations in Slovenian inflammatory bowel diseases patients
Authors:ID Mitrovič, Mitja (Author)
ID Potočnik, Uroš (Author)
Files:.pdf Disease_Markers_2011_Mitrovic,_Potocnik_High-resolution_melting_curve_analysis_for_high-throughput_genotyping_of_NOD2CARD15_mutations_an.pdf (1,72 MB)
MD5: 4614CA726E6206C26EA58740E6F0B82C
 
URL https://www.hindawi.com/journals/dm/2011/209101/abs/
 
Language:English
Work type:Scientific work
Typology:1.01 - Original Scientific Article
Organization:FKKT - Faculty of Chemistry and Chemical Engineering
Abstract:Inflammatory bowel diseases (IBD) are usually classified into Crohn's disease (CD) and ulcerative colitis (UC). NOD2/CARD15 was the first identified CD-susceptibility gene and was confirmed as the most potent disease gene in CD pathogenesis. Three NOD2/CARD15 variants, namely two missense polymorphisms R702W (rs2066844) and G908R (rs2066845), and a frame shift polymorphism L1007fs (rs2066847), were associated with CD in Caucasian populations. High resolution melting analysis (HRMA) with saturation LCGreen dyes was previously reported as a simple, inexpensive, accurate and sensitive method for genotyping and/or scanning of rare variants. For this reasons we used qPCR-HRMA for genotyping NOD2/CARD15 variants in 588 Slovenian IBD patients and 256 healthy controls. PCR-RFLP was used as a reference method for genotyping of clinical samples. The optimization of an HRM experiment required careful design and adjustment of main parameters, such as primer concentration, MgCl_{2} concentration, probe design and template DNA concentration. Different HRMA approaches were tested and used to develop a reliable and low-cost SNP genotyping assays for polymorphisms in NOD2/CARD15 gene. Direct HRMA was the fastest and cheapest HRMA approach for L1007fs and R702W polymorphisms, yet for G908R polymorphism sufficient reliability was achieved after introduction of unlabeled probe. In association analysis, we found statistically significant association of L1007fs (p =0.001, OR=3.011, CI95%=1.494-6.071) and G908R (p=2.62 * 10^{-4}, OR=14.117, CI95%= 1.884-105.799) polymorphisms with CD patients. At least one of NOD2/CARD15 polymorphisms was found in 78/354 (22.03% (12.69%) in UC patients and in 26/256 (10.15%) in healthy controls. We have successfully implemented NOD2/CARD15 HRMA assays, which may contribute to the development of genetic profiles for risk prediction of developing CD and for differential diagnosis of CD vs. UC.
Keywords:high-resolution melting analysis, NOD2/CARD15, inflammatory bowel diseases
Publication status:Published
Publication version:Version of Record
Year of publishing:2011
Number of pages:str. 265-274
Numbering:Letn. 30, št. 5
PID:20.500.12556/DKUM-66185 New window
ISSN:0278-0240
UDC:616.3
ISSN on article:0278-0240
COBISS.SI-ID:15127062 New window
DOI:10.3233/DMA-2011-0783 New window
NUK URN:URN:SI:UM:DK:5CDAT9GI
Publication date in DKUM:14.06.2017
Views:1516
Downloads:354
Metadata:XML DC-XML DC-RDF
Categories:Misc.
:
MITROVIČ, Mitja and POTOČNIK, Uroš, 2011, High-resolution melting curve analysis for high-throughput genotyping of NOD2/CARD15 mutations and distribution of these mutations in Slovenian inflammatory bowel diseases patients. Disease Markers [online]. 2011. Vol. 30, no. 5, p. 265–274. [Accessed 5 April 2025]. DOI 10.3233/DMA-2011-0783. Retrieved from: https://dk.um.si/IzpisGradiva.php?lang=eng&id=66185
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Record is a part of a journal

Title:Disease Markers
Shortened title:Dis. Markers
Publisher:Hindawi Publishing Corporation
ISSN:0278-0240
COBISS.SI-ID:1533717 New window

Document is financed by a project

Funder:ARRS - Slovenian Research Agency
Project number:J3-2175
Name:Gentski dejavniki tveganja in farmakogenomika kronično vnetne črevesne bolezni

Licences

License:CC BY 4.0, Creative Commons Attribution 4.0 International
Link:http://creativecommons.org/licenses/by/4.0/
Description:This is the standard Creative Commons license that gives others maximum freedom to do what they want with the work as long as they credit the author.
Licensing start date:14.06.2017

Secondary language

Language:Slovenian
Keywords:kronično vnetna črevesna bolezen


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