Published online before print September 20, 2007

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Copyright © 2007 American Society for Investigative Pathology
Journal of Molecular Diagnostics, doi:10.2353/jmoldx.2007.070027

Accepted for publication August 8, 2007.

Article

Simultaneous Amplification, Detection, and Analysis of Common Mutations in the Galactose-1-Phosphate Uridyl Transferase Gene

Mohamed Jama^*, Lesa Nelson, Rong Mao^*, and Elaine Lyon^*@

From the ARUP Institute for Clinical and Experimental Pathology,* Salt Lake City; Axial Biotech, Inc., Salt Lake City; and the Department of Pathology, University of Utah, Salt Lake City, Utah

^@ To whom correspondence should be addressed. E-mail: lyone{at}aruplab.com.

	Abstract

Classic galactosemia is an autosomal recessive inherited error of galactose metabolism. It is caused by lack of galactose-1-phosphate uridyl transferase, an enzyme that is required to metabolize galactose-1-phosphate to uridine diphosphate galactose. The build up of galactose-1-phosphate is toxic at high levels and can damage the liver, brain, eyes, and other vital organs. Over 200 mutations have been identified in affected individuals. We describe an assay to identify nine target mutations or variants in the galactose-1-phosphate uridyl transferase gene, namely p.Q188R, p.S135L, p.K285N, p.L195P, p.T138M, p.Y209C, IVS2-2 A>G, p.L218L, and p.N314D. A single long-range PCR is followed by a multiplexed nucleotide extension assay (single nucleotide extension) and capillary electrophoresis to detect simultaneously all nine target mutations/variants. Fifty-four previously characterized samples (47 clinical samples and seven controls) gave a 100% concordance. We also report a nontarget novel mutation, p.L192X, and its profile using single nucleotide extension. This assay can complement the enzyme activity assay and identify familial mutations for testing additional family members.