RESULTS
Two experimental approaches were utilized to study the iri vitro mito
chondrial RNA synthesis in Euglena gracilis. The first approach was to
study the incorporation of radioactively labeled ribonucleoside triphos
phates into acid insoluble products by isolated mitochondria which had
been treated with isotonic buffer to assure swelling and permability.
The products of the incorporations were then studied. The second approach
was to study the activity of solubilized and partially purified mitochon
drial DNA dependent RNA polymerase. This approach involved determining
the conditions for solubilizing the enzyme and then for partially purifying
the enzyme. The activity of this enzyme was then characterized.
The Incorporation of Ribonucleoside
Triphosphates by Isolated Mitochondria
The initial research objectives were to determine if RNA synthesizing
activity is retained by mitochondria isolated from Euglena gracilis, and
to look for ways to differentiate between nuclear and mitochondrial RNA
polymerase activities such as sensitivities to inhibitors or special
enzymatic requirements. The approach was to study the RNA synthesizing
activity of isolated mitochondria obtained from a streptomycin bleached
aplastidic mutant of Euglena gracilis, strain z. The mitochondrial
isolation procedure (Figure 1) consisted of rupturing washed cells in the
French pressure cell at 1000 lb/in^ and collecting crude mitochondrial
34