title: There exists a distinct stage during mammalian DNA synthesis immediately after joining of replication intermediates.
abstract: We describe an approach, using alkaline cell lysis and digestion with nuclease S1, which permits to distinguish between newly ligated DNA and the DNA of mature chromatin. When cells with steady-state labelled DNA (mature DNA) are analyzed, the results show labelled "nucleosomal-sized" DNA. However, when DNA of cells pulse-labelled with thymidine for 45 seconds is examined one can detect only large DNA. The newly ligated DNA is not reduced to "nucleosomal-sized" DNA by nuclease S1. When the large DNA is denatured in formamide one can detect 10 kb DNA fragments. Furthermore in pulse-chase experiments there appear, after formamide-treatment, increasing amounts of "nucleosomal-sized" DNA with a parallel decrease in the amount of 10 kb DNA fragments. Hence the newly ligated, large, DNA differs from mature DNA and represents a distinct stage during DNA replication.
mesh_terms: D002478:Cells, Cultured; D002843:Chromatin / Q000648:ultrastructure; D004261:DNA Replication; D004720:Endonucleases; D005559:Formamides; D006801:Humans; D008545:Melanoma; D008836:Micrococcal Nuclease; D009707:Nucleosomes / Q000648:ultrastructure*; D015719:Single-Strand Specific DNA and RNA Endonucleases; D014965:X-Rays
chemical_list: D002843:Chromatin; D005559:Formamides; D009707:Nucleosomes; C031066:formamide; D004720:Endonucleases; D015719:Single-Strand Specific DNA and RNA Endonucleases; D008836:Micrococcal Nuclease