REPLI-g Mitochondrial DNA Kit (25)

A whole-genome amplification solution from QIAGEN, the REPLI-g Mitochondrial DNA Kit enables selective amplification of mitochondrial DNA directly from total DNA samples, eliminating the need for time-consuming prior mitochondrial DNA isolation. Built on Multiple Displacement Amplification (MDA) technology, the kit uses a uniquely processive DNA polymerase that performs isothermal genome amplification and is capable of amplifying up to 100 kb without dissociating from the template, minimizing unequal sequence and locus representation compared with PCR-based methods. The procedure enriches for mitochondrial DNA with minimal contamination from nuclear DNA.

The workflow uses a heat denaturation step with a maximum input volume of 10 µl template DNA and a minimum pipetting volume of 1 µl, in a 50 µl reaction that runs for approximately 8 hours (overnight) at Mid throughput. It is designed for use with ~10 ng of purified total human genomic DNA and produces a yield of ~4 µg of amplified mitochondrial DNA, representing approximately 40 million-fold amplification. Due to the very high copy numbers per microliter of amplified product, the manufacturer recommends diluting the amplified DNA 1:1000 with nuclease-free water and using 1 µl of the diluted sample per PCR reaction.

The amplified DNA is suitable for downstream applications including genotyping, sequencing, and RFLP analysis. While optimized for human mitochondrial DNA, the kit can also be used for non-human species when the REPLI-g Human mt Primer Mix is substituted with an appropriate species-specific primer mix, which is not provided with the kit. Intact mitochondrial DNA is required for successful amplification.