A rapid and efficient method for enriching mitochondrial DNA from plants

Abstract

Current mitochondrial purification techniques are tedious and protracted due to their emphasis on recovering physiologically active mitochondria. However, for studies that are exclusively interested in isolating mitochondrial DNA (mtDNA) for applications such as PCR and sequencing, respiring mitochondria − and the complex procedures that stem from the need to retain their function − are unnecessary. Still, global DNA extraction methods have proven insufficient for mitochondrial DNA isolation because nuclear mitochondrial DNA segments (NUMTs) pose unique challenges to accurate mtDNA quantification and characterization. We present a rapid and simple extraction technique that maximizes recovery of mitochondrial DNA from plant cells, while minimizing the presence of nuclear DNA. Through real-time PCR, we show that this method provides a significant increase in the enrichment of mitochondrial DNA compared to that of nuclear DNA in both Arabidopsis thaliana and Brassica rapa. This method has important implications for future mitochondrial DNA analyses as it possesses few procedural limitations and minimizes the analytical problems typically associated with mtDNA purification by other techniques.