Prevention of a molecular misdiagnosis in galactosemia

Purpose: The polymerase chain reaction is generally used for mutational analysis of the galactose-1-phosphate uridyl transferase (GALT) gene in the diagnosis of galactosemia. This method is problematic when used in families of Ashkenazi Jewish descent. Methods: We amplified the GALT gene from leukocyte DNA followed by allele specific oligonucleotide hybridization, DNA sequencing and Southern Blot analysis to determine the mutant alleles causing galactosemia in a representative Jewish family. Results: The proband's diagnosis of galactosemia was confirmed by high levels of erythrocyte galactose-1-phosphate, absence of erythrocyte GALT activity and impaired total body oxidation of galactose to expired CO2. Initial molecular analysis of GALT alleles in the family showed homozygosity for a K285N missense mutation in the proband, homozygosity for N314D in the mother and heterozygosity for N314D and K285N in the father. These results contradicted Mendelian logic. Southern blot hybridization with GALT cDNA proved the presence of a complex 5 kb GALT deletion in the proband and her mother's DNA enabling a corrected genotype. Conclusions: Since a deletion of the GALT gene is a common mutation causing galactosemia among Ashkenazim Jewish families, this deletion should be suspected and tested for by genomic hybridization or by using primers specific for the 5 kb deletion.