In vitro rabbit mammary gland chromatin replication studied by micrococcal nuclease digestion.

Isolated nuclei from pregnant rabbit mammary glands were labelled with [3H]dTTP under conditions for DNA synthesis and subsequently digested with micrococcal nuclease. Replicating chromatin was found to exhibit increased susceptibility towards the nuclease. Analysis of chromatin digestion products by sucrose density gradient centrifugation demonstrated the association of in vitro replicated DNA with nucleosomes. Furthermore, the distribution of DNA polymerizing activity was studied in isolated nuclease-digested mammary gland chromatin. About 90% of all recovered nuclear DNA polymerizing activity cosedimented with nucleosomal particles, mainly with mononucleosomes. The distribution of DNA polymerases alpha and beta in chromatin isolated from the mammary glands of pregnant and lactating rabbits was compared. In these physiological states the mononucleosome-associated DNA polymerase alpha activity varied in accordance with the rate of DNA synthesis.