The first step is addition of lysis buffer to the samples. Cell nuclei and mitochondria are pelleted by centrifugation and resuspended in denaturation buffer containing QIAGEN Protease. Following protein digestion, DNA is precipitated by addition of isopropanol, recovered by centrifugation, washed in 70% ethanol, and dried. DNA is resuspended in hydration buffer and is ready for direct use in downstream assays or storage at –20ºC.
Variable amounts of DNA template were used to amplify the single-copy Hugl gene and a mitochondrial target (tRNAlys/ATPase). Each sample was analyzed 6 times, and reproducible results were achieved. M: markers.
Genomic DNA isolated from whole blood taken from 4 donors. Starting blood volumes are indicated above the lanes. C: blood treated with sodium citrate; E: blood treated with EDTA; M: markers.