DNA replication in homogenates of Physarum polycephalum

Homogenates of Physarum polycephalum incorporate [3H]dATP into nuclear DNA at an initial rate of approximately 15% of the in vivo rate. To attain this level of synthesis, cultures, are homogenized in a medium containing Mg++, EGTA, glucose and spermine. Incorporation is strongly stimulated by the addition of ATP and all four deoxyribonucleoside triphosphates to homogenates prior to incubation. Various inorganic cations other than Mg++ either do not affect synthesis or are inhibitory. Incorporation is inhibited by a nonionic detergent, Triton X-100. DNA synthesis in this cell-free nuclear system is similar in several respects to that which occurs in vivo: (1) The rate of DNA synthesis in the intact organism at a given time in the mitotic cycle is reflected by the level of synthesis in homogenates prepared from cultures at that time of the cycle; (2) DNA strands labeled in vitro exhibit alkaline sucrose density gradient sedimentation properties similar to those of daughter-strand DNA pulse-labeled in vivo; and (3) Homogenates of cultures which were pre-treated with cycloheximide incorporate [3H]dATP at about 60% of the level observed in homogenates of untreated controls.