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Figure 4 Quantification and distribution of the A4300G mutation in family 2. (A) Labeled products generated by the same restriction fragment length polymorphism (RFLP) analysis described in Figure 3, showing the homoplasmic mutation in the heart deoxyribonucleic acid of both probands (IV-03 in family 1 and IV-01 in family 2) and in the fibroblasts of the proband (IV-01) from family 2. Fragment sizes (bp) are shown on the right. (B) Scheme of the alternative RFLP analysis first described by Casali et al. (14). In combination with the A4300G mutation, a mismatched forward primer creates a new HphI recognition site that cuts the 263-bp polymerase chain reaction-amplified product into two smaller products of 235 and 28 bp. In this assay, wild-type mitochondrial deoxyribonucleic acid (mtDNA) remains uncut. (C) Representative gel of samples from three family members showing labeled products generated by the RFLP analysis described earlier. A total of 23 maternal relatives were screened by this assay and shown to be homoplasmic for the A4300G mutation. Fragment sizes (bp) are shown on the right.