NO EFFECT OF TREATMENT WITH CARCINOGENS ON CYTOSINE METHYLATION OF MITOCHONDRIAL-DNA ISOLATED FROM RAT ORGANS BY PHENOL-FREE ALKALINE EXTRACTION

Due to the postulated role of mitochondrial DNA damage in aging as wel l as in the pathogenesis of several chronic degenerative diseases, inc luding tumors, there is a need for easy, safe, rapid and inexpensive m ethods for mitochondrial DNA isolation, We propose a simple, 1-day pro tocol based on alkaline extraction, which avoids time-consuming gradie nt centrifugations and use of toxic phenol. This procedure was used fo r the recovery of mitochondrial DNA from different rat organs (liver, kidney, heart, lung, brain and testis). The yield was quite high. Puri ty was sufficient enough to perform restriction analyses with several endonucleases. No changes were observed in the structure or methylatio n patterns of -CCGG- sites in liver mitochondrial DNA isolated from ra ts exposed whole-body to mainstream cigarette smoke or treated with th e carcinogens benzo[a]pyrene, 2-acetylaminofluorene or diethylnitrosam ine.