Isolation, fractionation, and RNA polymerase activity in a sample enriched with oligodendroglia nuclei

RNA polymerase activity in human brain tumors

“…Nuclear Isolation. The nuclear isolation procedure was similar to that described previously (Slagel and Akers, 1972). The entire procedure was carried out at 0--4~ The tumor tissue was minced and homogenized as a 10~ suspension in a glass homogenizer with a teflon pestle having a 0.01 inch clearance.…”

“…Dravid et al, (1969) have demonstrated RNA polymerase activity in isolated brain tumor nuelei. Because of the restraints placed on the experiment by the small quantity of tissue from human tumor biopsies, we have characterized the in vitro RNA polymerase activity in isolated normal sheep glial cell nuclei with respeet to pI-I optimum, time course of the reaction and ammonium sulfate activation (Slagel and Akers, 1972). It was also determined that 80 to 900/0 of the in vitro activity is due to an alpha-amanitin inhibitable enzyme (Slagel, unpublished).…”

Effect of GTP and Other Nucleoside Triphosphates on GMP Incorporation by Isolated Cortical Nuclei

“…vitro by brain nuclei has extendeifover a wide range; this is particularly true for thelabelei nucleoside triphosphate (Slagel & Akers, 1972;Banks & Johnson, 1973;, Austoker et~, 1972). The effect of this wide range of substrate concentration has not been previously reported.…”

“…All these authors used relatively high concentrations of the labeled nucleoside triphosphate .. Slagel and Akers (1972) and Banks and Johnson (1973), using a low concentration of GTP, reported a lower optimal ammonium sulfate concentration. We found at low GTP concentration in the standard -14-medium that the stimulation by ammonium sulfate was maximal at 50 rnt1.…”

“…The only other reported inhibition by actinomycin at 10\." GTP concentration and high ionic strength was in the presence of 400 pg/ml of the inhibitor (Slagel and Akers, 1972) *Preincubations were at 37°C f.or 10 min. The preincubation volume was 480 ~1 in tubes with final 0.71 pM GTP.…”