Induction and Repair of Double- and Single-strand DNA Breaks in Bacteriophage Lambda SuperinfectingEscherichia Coli

Abstract

Induction and repair of double- and single-strand DNA breaks were measured after decays of 125I and 3H incorporated into the DNA and after external irradiation with 4 MeV electrons. For the decay experiments, cells of wild type E. coli K-12 were superinfected with bacteriophage .lambda. DNA labeled with 5''-(125I)iodo-2''-deoxyuridine or with (methyl-3H)thymidine and frozen in liquid N. Aliquots were thawed at intervals and lysed at neutral pH, and the phage DNA was assayed for double- and single-strand breakage by neutral sucrose gradient centrifugation. The gradients used allowed measurements of both kinds of breaks in the same gradient. Decays of 125I induced 0.39 single-strand breaks/double-strand break. No repair of either break type could be detected. Each 3H disintegration caused 0.20 single-strand breaks and very few double-strand breaks. The single-strand breaks were rapidly rejoined after the cells were thawed. For irradiation with 4 MeV electrons, cells of wild type E. coli K-12 were superinfected with phage .lambda. and suspended in growth medium. Irradiation induced 42 single-strand breaks/double-strand break. The rates of break induction were 6.75 .times. 10-14 (double-strand breaks) and 2.82 .times. 10-12 (single-strand breaks)/rad and per dalton. The single-strand breaks were rapidly repaired upon incubation whereas the double-strand breaks seemed to remain unrepaired. Double-strand breaks in superinfecting bacteriophage .lambda. DNA are repaired to a very small extent, if at all.