A3243G mutation in the mitochondrial genome is the most frequent mutation of mitochondrial diseases in humans and the major cause of mitochondrial encephalomyopathy lactic acidosis, stroke like episodes (MELAS). The A3243G mutation exists in the heteroplasmic form (heteroplasmy) and the severity of the diseases is correlated with the percentage of the mitochondrial genome copies which carry the mutation. In this study, the RFLP-PCR was used in combination with real-time PCR using Taqman LNA (locked nucleic acid) fluorescent probes to detect and quantitate the percentage of the A3243G mutation. According to the PCRRFLP analysis, the normal subjects have only 198 bp amplified band while the patients have the 198 bp band (of the unmutated copies) and III bp and plus 87 bp bands (of the mutated copies). Eighty two patients with the encephalopathy symptoms were screened for the presence of the A3243G mutation and 4 patients (3 male and 1 female, 11-15 years old) were found to carry the A3243G mutation. The A3243G mutation was also detected in their mother and siblings but not in their father of the patients. The obtained results were reconfirmed by DNA sequencing. It was also found in the PCR-RFLP banding patterns that the mother of the patients had rather faint and tiny bands of 111 bp and 87 bp compared to the 198 bp band. Using real-time PCR with Taqrnan LNA fluorescent probes, the MELAS patient of family II was shown to carry 73.02 percent of the A3243G-carrying genome copies compared to 9.24 percent in his mother. This finding was in good agreement with the PCR-RFLP banding pattern and explained why his mother did not show any symptom of the disease.
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