DETECTION AND ANALYSIS OF MITOCHONDRIAL-DNA DELETIONS BY WHOLE GENOMEPCR

Southern blot analysis has been the best method available for the scre ening and detection of mitochondrial DNA (mtDNA) rearrangements. Recen t developments in polymerase chain reaction (PCR) technology allowed t he amplification of the whole mitochondrial genome (16.6 kb), making P CR a potentially useful technique for the detection of mtDNA deletions . We tested the usefulness of whole mitochondrial genome PCR by studyi ng skeletal muscle DNA from seven patients with single and multiple de letions and controls from ages 3 to 91 years old. Specific patterns fo r single and multiple deletions were obtained with whole genome PCR, w hich were confirmed by the Southern analysis with probes hybridizing t o mtDNA sequences. Amplifications from young controls (3 to 23 years o ld) yielded only one band (16.5 kb) while amplification from older con trols revealed one or more additional smaller bands. The amplification from the 91-year-old control showed a pattern similar to amplificatio ns from patients with multiple mtDNA deletions. Although single and mu ltiple mtDNA deletions could be readily detected from patient samples, the high sensitivity of this method can lead to false positive result s due to the presence of age-related deletions in old control samples. Despite its limitations, whole mitochondrial genome PCR can be useful for the detection of single deletions in muscle samples obtained from young individuals since the levels of age-related deletions are too l ow to be amplified. (C) 1996 Academic Press, Inc.