Detection Of Homoplasmic And Heteroplasmic Mutations In Mitochondrial Genome

Energy metabolism disorders caused by mutations in mtDNA represents a group of heterogeneous diseases. The most frequently found mtDNA point mutations are A3243G for mitochondrial myopathy, encephalopathy, lactic acidosis, and stroke-like episodes (MELAS), A8344G for myoclonus epilepsy, ragged red fibers (MERRF), T8993G/C for neuropathy, ataxia, retinitis pigmentosa (NARP), and Leigh syndrome, and G11778A for Leber's hereditary optic neuropathy (LHON).[16,22] Molecular analysis of the known mtDNA mutations is usually carried out by polymerase chain reaction/ restriction fragment length polymorphism (PCR/RFLP) or multiplex PCR/ASO (allele-specific oligonucleotides) dot blot (cross reference) method. Recently, a capillary-based technique was also used for A3243G mutation analysis.[23] Using multiplex PCR/ASO method to screen 44 known point mutations in 2000 patients with mtDNA disorders, we have demonstrated that only about 6% of the patients had identifiable, disease-causing mutations.[16,22] These data suggest that the majority of the pathogenic mutations have yet to be identified.

One of the unique characteristics of mitochondrial genetics is heteroplasmy. Because there are hundreds to thousands of mitochondria per animal cell, the percentage of mutant mitochondria can vary from 0% to 100%, unlike the nuclear gene mutation that is 100% for homozygous and 50% for heterozygous. In addition, there are numerous homoplasmic mtDNA variations. Thus the requirement for an effective mutation detection method is the ability to distinguish between the heteroplasmic and homoplasmic mtDNA mutations.

Both psoralen clamp and GC clamp have been used for broad-range (0-80% denaturant) denaturing gradient gel electrophoresis (DGGE).[24,25] Michikawa et al.[25] developed the DGGE conditions for 18 psoralen-clamped DNA segments to analyze 20 different known mtDNA point mutations in tRNA genes. Sternberg et al.[24] used the traditional DGGE method with a GC clamp to investigate 35 unrelated patients with mitochondrial

Getting Started With Dumbbells

Getting Started With Dumbbells

The use of dumbbells gives you a much more comprehensive strengthening effect because the workout engages your stabilizer muscles, in addition to the muscle you may be pin-pointing. Without all of the belts and artificial stabilizers of a machine, you also engage your core muscles, which are your body's natural stabilizers.

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