Flow cytometric evaluation of DNA digestion with S1 nuclease.
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.
Prosperi
E
E
Dipartimento di Biologia Animale dell'Università, Pavia, Italy.
Bottiroli
G
G
Supino
R
R
D000818:Animals; D002460:Cell Line; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism*; D004273:DNA, Neoplasm / Q000187:drug effects / Q000378:metabolism* / Q000528:radiation effects; D004720:Endonucleases / Q000494:pharmacology*; D005434:Flow Cytometry; D006801:Humans; D008214:Lymphocytes / Q000187:drug effects / Q000378:metabolism*; D051379:Mice; D008810:Mice, Inbred C57BL; D009691:Nucleic Acid Denaturation / Q000187:drug effects*; D015719:Single-Strand Specific DNA and RNA Endonucleases; D013154:Spleen / Q000166:cytology / Q000187:drug effects; D013997:Time Factors
D004273:DNA, Neoplasm; D004720:Endonucleases; D015719:Single-Strand Specific DNA and RNA Endonucleases