Thus, it is notclear whe ther the concentra tion of salt used in situ, the agentused to damage DNA, or thecell type employed was responsble for theconflicting repor ts. Th is communication presen ts preliminary resuits in vestigating two specific variables: the concentration of salt employed in situ, and the ag entusedto induce repa ir syn thes is. For DNA <a href="https://www.ncbi.nlm.nih.gov/pubmed/19485832">buy
Doxycycline hyclate</a> repair synthesis, GI cells were treated with MNNG or NMU before permeabilization or with neocarzinostatin or bleomycin after permeabilization.Permeable GI cells were incubated in the solution described for DNA replication, except TTP was. mM in all repair experiments except those utilizing neocarzinostatin, where mM dithiothreitol was added.Aphidicolin and araCTP inh ibit polymerase a, whe reas ddTTP inh ibitspolymerase. At mM KC, p araCT P or pgml aphidicolin inh ibited replication or, respectively, while pdd T T P only reduced replica tion. The se concentrations of inh ibitors produced maximum inhibition.Essentially identical results were ob ta ined wi th TTP reduced to the concen tration used for repa ir syn thes is. mM incubated with in tactcells for hat C in low isoleucine medium before permeabilization; NMU, mM incubated with intact cells for hat C in low isoleucine medium before permeabilization.. or pgml aphidicolin inhibited replication or, re spectively, whereas p ddTTP reduced replication. DNA repair synthesis was induced in GI cells by a variety of agents.In the absence of repairinducing agents, permeable G I cells incorporated little TTP into DNA. A higher concentration of dithiothre itol. The DNA synthe sued in response to neocarzinostatin banded atnormal den sity, which is characteristic of repair synthesis. Neocarzinostatin repair synthesis was higher at mM KC than in the absence of KC. MNNG and NMU induced more DNA synthesis when prein cubated with intact cells before permeabilization than when incubated directly with permeable cells.BrdUTP densitylabeled DNA synthesized in response to both alkyl ating agents banded atnormal DNA density, indicating in duction of DNA repair synthesis. Repair synthe sis induced by MNNG or NMU was higher in the absence of KC than in the presence of KC. Bleomycin and neocarzinostatin repair synthesis was reduced by the polymerasepinhibitor, ddTTP. In contrast, the polymerase a inhibitors had little effect on bleomycin repair, although aphidicolin significantly re duced neocarzinostatin repair. Inhibition of bleo mycin or neocarzinostatin repair synthesis by ddTTP aphi dicolin was somewhat greater than thatby ddTTP alone. Higher concentrations of the polymerase inhibitors did not significantly increase the inhibition of repair synthesis.These findings indicate thatpolymerasepparticipated to a greater extent than polymerase a in bleomycin and neocarzi P. We then determined whether low KC altered the effect of DNA polymerase inhibitors on repair synthesis.Low salt did not appreciably alter the effect of ddTTP on DNA repair synthesis. However, the sensitivity of repair syn thesis to inhibition by the polymerase a inhibitors was in creased for most DNA damaging agents atlow KC. The inhibition of MNNG and NMU repair synthesis by araCTP was especially pronounced atlow KC.The amount of DNA damage induced by each agent was investigated.The effect of polymerase inhibitors is presented as percentage of inhibition of repair synthesis in control cells.