Philippe Joly1, Philippe Lacan, Caroline Garcia, Cyril Martin, Alain Francina. 1. Unité de Pathologie Moléculaire du Globule Rouge, Fédération de Biochimie et de Biologie Spécialisée, Hôpital Edouard Herriot, Hospices Civils and Université Claude Bernard-Lyon 1, Lyon, France.
Abstract
OBJECTIVES: The Mediterranean and A(-) G6PD variants are particularly prevalent in Africa and Southern Europe. Our study was aimed to develop an assay for the rapid genotyping of these two variants by HRM. METHODS: After PCR reactions corresponding to the G6PD Mediterranean (exon 6), G6PD (A-) (exon 4) and G6PD (A-) (exon 5) mutations, amplicons were submitted to HRM. This protocol was applied to a cohort of 132 patients suffering from sickle cell disease. RESULTS: Wild, homozygous or hemizygous and heterozygous states were fully discriminated by HRM for all three mutations. HRM results were in total accordance with DNA sequencing for 22 patients of our cohort with a 'A' genotype: presence of the (A-) (exon 5) mutation but absence of the (A-) (exon 4) mutation. CONCLUSIONS: Our HRM protocols allow a rapid, simple and cost-effective screening of G6PD deficiency in patients originating from the Mediterranean and the African areas. Copyright 2009 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.
OBJECTIVES: The Mediterranean and A(-) G6PD variants are particularly prevalent in Africa and Southern Europe. Our study was aimed to develop an assay for the rapid genotyping of these two variants by HRM. METHODS: After PCR reactions corresponding to the G6PD Mediterranean (exon 6), G6PD (A-) (exon 4) and G6PD (A-) (exon 5) mutations, amplicons were submitted to HRM. This protocol was applied to a cohort of 132 patients suffering from sickle cell disease. RESULTS: Wild, homozygous or hemizygous and heterozygous states were fully discriminated by HRM for all three mutations. HRM results were in total accordance with DNA sequencing for 22 patients of our cohort with a 'A' genotype: presence of the (A-) (exon 5) mutation but absence of the (A-) (exon 4) mutation. CONCLUSIONS: Our HRM protocols allow a rapid, simple and cost-effective screening of G6PD deficiency in patients originating from the Mediterranean and the African areas. Copyright 2009 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.