The digestion of DNA with the S1 nuclease has been investigated by flow cytometry on two cultured cell lines and on splenic lymphocytes. The nucleolytic activity of the enzyme has been determined by measuring the decrease in fluorescence intensity of DNA stained with Propidium Iodide (PI). The reaction kinetics could be estimated through the shift of the G1 peak, since the shape of the DNA histograms obtained from cell submitted to S1 digestion did not show any particular variation as compared with the histograms of undigested control cells. DNA digestion was found to be more pronounced when cells were previously damaged with agents known to induce strand breaks, such as gamma-rays or Adriamycin. The assessment of DNA sensitivity to S1 nuclease with flow cytometry may thus provide a fast and reliable procedure for evaluating chromatin structural changes, both in physiological and pathological conditions.
Flow cytometric evaluation of DNA digestion with S1 nuclease.
Prosperi E;Bottiroli G;
1987
Abstract
The digestion of DNA with the S1 nuclease has been investigated by flow cytometry on two cultured cell lines and on splenic lymphocytes. The nucleolytic activity of the enzyme has been determined by measuring the decrease in fluorescence intensity of DNA stained with Propidium Iodide (PI). The reaction kinetics could be estimated through the shift of the G1 peak, since the shape of the DNA histograms obtained from cell submitted to S1 digestion did not show any particular variation as compared with the histograms of undigested control cells. DNA digestion was found to be more pronounced when cells were previously damaged with agents known to induce strand breaks, such as gamma-rays or Adriamycin. The assessment of DNA sensitivity to S1 nuclease with flow cytometry may thus provide a fast and reliable procedure for evaluating chromatin structural changes, both in physiological and pathological conditions.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
