High-throughput genotyping of mannose-binding lectin variants using high-resolution DNA-melting analysis

Human Mutation
Rolf H A M VossenAnja Roos

Abstract

High Resolution Melting Analysis (HRMA) is a rapid and sensitive method for single nucleotide polymorphism (SNP) analysis. In the present study we present a novel HRMA assay to detect three SNPs in close proximity of each other in the first exon of the gene encoding mannose-binding lectin (MBL), a key molecule of innate immunity. These SNPs have been selected for their known biological and clinical relevance. The three SNPs in MBL2 were simultaneously determined in sixty-nine human DNA samples using HRMA and a single non-fluorescent melting probe, without any post-PCR processing of samples. Combining analyses from amplicon melting and probe melting, we have been able to discriminate ten exon 1 MBL2 genotypes with HRMA, making it a suitable tool for MBL genotyping. A second HRMA assay is presented to detect a relevant polymorphism (Y/X SNP) in the MBL2 promoter region. In conclusion, HRMA is a closed tube assay that is easy to setup and lends itself perfectly for high throughput genotyping of MBL2 variants. The present study thereby facilitates further clinical studies into the role of MBL in inflammatory and infectious disease.

Citations

Oct 12, 2012·International Journal of Hematology·Xinju ZhangMing Guan
Jun 10, 2010·Allergy, Asthma, and Clinical Immunology : Official Journal of the Canadian Society of Allergy and Clinical Immunology·Rohan AmeratungaDonald R Love
Jul 13, 2013·International Journal of Biological Sciences·Chunming WangJianmin Wan
Jul 2, 2010·Hepatology : Official Journal of the American Association for the Study of Liver Diseases·Bert-Jan F de RooijHein W Verspaget
Nov 15, 2013·The British Journal of Surgery·M W G A BronkhorstE M M van Lieshout
Apr 10, 2012·Best Practice & Research. Clinical Gastroenterology·Bart van HoekHein W Verspaget
Feb 22, 2013·The Journal of Trauma and Acute Care Surgery·Maarten W G A BronkhorstEsther M M Van Lieshout
Nov 20, 2012·Electrophoresis·Masaru AsariKeiko Shimizu

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