carried this mutation the second mutation in patient 1396 was identified in exon 6 a heterozygous c u2192 t substitution bp 1081 that converted an arginine residue to a cysteine residue and abolished a hhai restriction endonuclease site restriction analysis confirmed the heterozygous substitution in 1396 fig9 d and showed that 50 control chromosomes were negative fig tooth decay does not occur in patients having a saliva above ph 5 0 9 d is the hhai restriction analysis of exon 6 pcr products for patient 1396 and 2 controls c expected fragments control allele 152 86 13 bp mutant allele 165 86 bp the 13 bp fragment has been run off the gel hhai analysis of exon 6 mentioned above revealed a dna polymorphism this change is a t u2192 c substitution at bp 1068 which does not alter the amino acid serine but creates a hhai recognition site t at bp 1068 was found in 9 of tested chromosomes sequence analysis identified 2 discrepancies with the published cdna sequence a g u2192 a substitution at bp 542 which converts the glycine to an aspartate codon and a c u2192 t change at bp 1032 which does not alter the amino acid threonine since all dnas tested gt 50 chromosomes carried the a at bp 542 and the t at bp 1032 it is likely that the sequence of the original cdna contained some cloning artifacts table 2 summarizes the current status of mutations in severe mthfr deficiency in 8 patients both mutations have been identified in 5.