The covalently closed circular duplex deoxyribonucleic acid (DNA) of øX174 underwent progressive conversion to nicked and linear DNA with increasing bleomycin øX174 RFI DNA molecule ratios. The formation of linear DNA (a double-strand break) occurred under limited reaction conditions as low as an average of 0.2 single-strand break øX174 RFI DNA molecule. As bleomycin-produced linear DNA was further fragmented by bleomycin, a broad distribution of DNA fragments without notable concentrations of unique size was formed. Restriction enzymes Pstl and Stoll did not generate discrete fragments from bleomycin-produced full-length linear øX174 DNA, nor did bleomycin cleavage generate discrete fragments from Hpall or Pstl digests of øX174 RFI. These findings suggest that bleomycin does not act at a few specific sites on øX174 RFI DNA. The singlestrand nick appeared to be the preferred site for bleomycin action for a second cleavage in a øX174 molecule.
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