The main stages of the run-on transcription method as applied to the quantification of mitochondrial gene transcription are described. As distinct from widely applied methods of northern hybridization and PCR after reverse transcription, this method permits studying the effects of exogenous and endogenous factors on the rate of mitochondrial gene transcription. The main method stages are (1) isolation of intact organelles; (2) preparation of nylon membranes with gene-specific fragments of DNA; (3) transcription in the lysates of these organelles in the presence of 32P-labeled RNA precursors; and (4) hybridization of newly synthesized labeled transcripts with DNA fragments of tested mitochondrial genes loaded on the nylon membrane.
Russian Journal of Plant Physiology – Springer Journals
Published: Jan 8, 2011
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