An Improved Method to Isolate Mitochondrial RNA from Green Plant Tissue
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In plant cells, mitochondrial RNA (mtRNA) constitutes about only 1% of the total RNA. From this, most RNAs are ribosomal RNAs. Thus, isolation of high purified mtRNA is necessary, not only for construction of a mitochondrial cDNA library, but also for the analysis of plant mitochondrial transcription. Several methods have been frequently used for isolation of plant mtRNA (1, 2, 3). However, these mtRNA preparations may be heavily contaminated by chloroplast RNA (cpRNA), especially when mtRNA is isolated from green leaves (1,4). It is believed that the cpRNA sticks to the mitochondrial membrane and therefore persists after gradient purification of mitochondria. Although micrococcal nuclease would be the enzyme to remove the non-mtRNA from mitochondrial membranes prior to lysis of mitochondria, treatments with micrococcal nuclease for the mtRNA isolation from green leaves have not been effective (4).
KeywordsPotassium Acetate Micrococcal Nuclease Guanidinium Thiocyanate Sucrose Gradient Centrifugation Guanidine Thiocyanate
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