title: Detection of a 10 kb DNA replication intermediate in human melanoma cells.
abstract: DNA replication in human melanoma cells is investigated by lysing the cells in dilute alkali. This lysis condition results in the release from parental DNA of the single-stranded DNA fragments located in active replicating units. The size of the released DNA should theoretically range from that of Okazaki-fragments up to that of the entire replication unit. However, the results showed that the released DNA replication intermediates which are detected range in the size between Okazaki-fragments up to 10 kb DNA fragments. The 10 kb DNA fragments show a discrete appearance in agarose gel electrophoresis. Moreover the kinetic results indicate that the ligation of the 10 kb DNA fragments to form high molecular weight DNA is a late step. A prerequisite for the release of this DNA fragment as a discrete population is that there are gaps in the continuity of the newly synthesized DNA spaced roughly 10 kb away from each other.
mesh_terms: D002478:Cells, Cultured; D002499:Centrifugation, Density Gradient; D004247:DNA / Q000378:metabolism; D004261:DNA Replication; D004277:DNA, Single-Stranded / Q000378:metabolism; D004587:Electrophoresis, Agar Gel; D006801:Humans; D008545:Melanoma / Q000235:genetics; D012972:Sodium Hydroxide / Q000494:pharmacology; D013936:Thymidine / Q000378:metabolism; D013997:Time Factors
chemical_list: D004277:DNA, Single-Stranded; C022262:Okazaki fragments; D012972:Sodium Hydroxide; D004247:DNA; D013936:Thymidine