Mitochondria are crucial organelles within cells that produce energy. They primarily generate ATP (adenosine triphosphate) through cellular respiration, which is essential for various cellular functions. Mitochondria have a double-membrane structure and contain their own mitochondrial DNA (mtDNA), separate from the nuclear DNA. This mtDNA is circular and double-stranded, consisting of approximately 16,569 base pairs in humans.
Mutations in the mitochondrial genome can impair energy production, leading to various diseases. Mitochondrial disorders often manifest in energy-demanding tissues such as the nervous system, muscles, heart, and auditory system. Additionally, somatic mutations in mitochondrial DNA have known to be linked to cancer and degenerative diseases, affecting disease occurrence and prognosis. Genetic testing is a valuable tool for accurate diagnosis of these conditions.
To achieve high-purity separation of mitochondrial DNA, complex centrifugation steps are required. However, there is a simpler approach that involves extracting total DNA from samples and then selectively amplifying mitochondrial DNA. Two common methods for mitochondrial DNA enrichment are PCR-based and hybridization-based methods.
The PCR-based method involves designing primer pairs that bind to the mitochondrial genome to amplify only the mitochondrial DNA. If the size of the amplicon becomes too large, amplification efficiency decreases, so the mitochondrial genome is divided into appropriate-sized segments for effective amplification. While PCR-based mitochondrial amplification is straightforward, it can be hindered by mutations at primer binding sites and pose limited scalability for analyzing mitochondrial DNA alongside other genomic regions.
The hybridization-based method uses multiple probes that are complementary to the mitochondrial genome to capture mitochondrial DNA. Although this method is more complex than PCR, it is less affected by mutations and provides more stable sequence information. It also allows for easy expansion to analyze other genomic regions by adding different set of probes.
Both methods have their advantages and limitations. For cases requiring single mitochondrial analysis and rapid processing of large numbers of samples, the simpler PCR method is more effective. Conversely, for comprehensive analysis of mitochondrial DNA alongside other genomic regions, the hybridization method is more suitable due to its scalability.
Celemics Mitochondrial DNA Panel: https://www.celemics.com/products/ready-to-use-ngs-panel/mitochondrial-dna/
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