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1
Establissement Francais du Sang-Bretagne,
2
Université de Bretagne Occidentale, and
3
Laboratoire de Génétique Moléculaire, INSERM (EMI-01 15), 46 Rue Félix Le Dantec, 29200 Brest, France.
aAddress correspondence to this author at: EFS-Bretagne, 46 Rue Félix Le Dantec, 29200 Brest, France. E-mail gerald.legac{at}univ-brest.fr.
Background: Between 4% and 35% of hereditary hemochromatosis (HC) probands are C282Y or H63D heterozygotes or lack both of these two common HFE mutations, and 15 novel HFE mutations have been described recently. We evaluated denaturing HPLC (DHPLC) for screening of the whole HFE coding region and further defined whether HC probands with an incomplete HFE genotype carry uncommon mutations.
Methods: Analytical conditions for each coding exon were determined by a combination of computer melting profile predictions and experimental melting curves. To test accuracy for scanning the complete HFE coding region and optimize DHPLC running conditions, each melting domain was investigated with at least one mutation or one polymorphism as reference. We tested 100 DNA samples harboring the C282Y, H63D, or S65C mutations and 17 artificially created positive controls that carried either 1 of the 14 other known HFE mutations or 3 selected polymorphisms.
Results: Investigations on each of the coding exons 1, 2, 4, 5, and 6 could be performed at one analysis temperature. Coding exon 3 displayed a more complex melting profile and required two analysis temperatures. DHPLC detected all known HFE mutations as well as the three selected polymorphisms.
Conclusions: DHPLC can be used to scan the HFE gene in HC probands in whom at least one chromosome lacks an assigned mutation.
The following articles in journals at HighWire Press have cited this article:
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  S. Jacolot, G. Le Gac, V. Scotet, I. Quere, C. Mura, and C. Ferec HAMP as a modifier gene that increases the phenotypic expression of the HFE pC282Y homozygous genotype Blood, April 1, 2004; 103(7): 2835 - 2840. [Abstract] [Full Text] [PDF]
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 G. Biasiotto, S. Belloli, G. Ruggeri, I. Zanella, G. Gerardi, M. Corrado, E. Gobbi, A. Albertini, and P. Arosio Identification of New Mutations of the HFE, Hepcidin, and Transferrin Receptor 2 Genes by Denaturing HPLC Analysis of Individuals with Biochemical Indications of Iron Overload Clin. Chem., December 1, 2003; 49(12): 1981 - 1988. [Abstract] [Full Text] [PDF]
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 V. Scotet, M.-C. Merour, A.-Y. Mercier, B. Chanu, T. Le Faou, O. Raguenes, G. Le Gac, C. Mura, J.-B. Nousbaum, and C. Ferec Hereditary Hemochromatosis: Effect of Excessive Alcohol Consumption on Disease Expression in Patients Homozygous for the C282Y Mutation Am. J. Epidemiol., July 15, 2003; 158(2): 129 - 134. [Abstract] [Full Text] [PDF]
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S. Fruchon, M. Bensaid, N. Borot, M.-P. Roth, and H. Coppin Use of Denaturing HPLC and a Heteroduplex Generator to Detect the HFE C282Y Mutation Associated with Genetic Hemochromatosis Clin. Chem., May 1, 2003; 49(5): 822 - 824. [Full Text] [PDF]
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O. A. Bodamer, D. Bercovich, M. Schlabach, C. Ballantyne, D. Zoch, and A. L. Beaudet Use of Denaturing HPLC to Provide Efficient Detection of Mutations Causing Familial Hypercholesterolemia Clin. Chem., November 1, 2002; 48(11): 1913 - 1918. [Abstract] [Full Text] [PDF]
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S. Pissard, L.-T.-A. Huynh, J. Martin, and M. Goossens HFE Genotyping by Amplification Refractory Mutation System-Denaturing HPLC Clin. Chem., May 1, 2002; 48(5): 769 - 772. [Full Text] [PDF]
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