Massively parallel sequencing (MPS) allows laboratories access to mitochondrial DNA (mtDNA) analysis using a more sensitive and high-throughput workflow compared to traditional sequencing methods. Increased mixture deconvolution and heteroplasmy resolution are achieved by deep sequencing coverage and digital read counts. Additionally, the use of small amplicons to sequence the mitochondrial control region improves sequencing results from degraded samples.
The PowerSeq™ CRM Nested System, Custom, generates 10 small amplicons covering the control region of the mitochondrial genome in one multiplex. The targeted regions for amplification are designed to be in a range of 144–237bp to ensure optimal results from degraded samples. The system uses a nested amplification protocol that greatly reduces the number of steps and time required to produce libraries ready for sequencing. The protocol consists of a single PCR step to both amplify the target amplicons and incorporate indexed sequencing adapters.