title: Chromatin fractionation analysis of licensing factors in mammalian cells.
abstract: ORC, Cdc6, Cdt1, and MCM2-7 are replication-licensing factors, which play a central role in the once-per-cell cycle control of DNA replication. ORC, Cdc6, and Cdt1 collaborate to load MCM2-7 onto replication origins in order to license them for replication. MCM2-7 is a DNA helicase directly involved in DNA replication and dissociates from DNA as S phase progresses and each replicon is replicated. In the cell cycle, the loading of MCM2-7 is restricted during the end of mitosis and the G1 phase. Thus, the levels of chromatin-bound MCM2-7 and its loaders oscillate during the cell cycle. Chromatin association of these factors can be analyzed by separating a cell lysate into soluble and chromatin-enriched insoluble fractions in mammalian cells.
authors.affiliation: Graduate School of Life Science, University of Hyogo, Kamigori, Ako-gun, Hyogo, 678-1297, Japan, hideon@sci.u-hyogo.ac.jp.
authors.forename: Hideo
authors.initials: H
authors.lastname: Nishitani
authors.forename: Masayuki
authors.initials: M
authors.lastname: Morino
authors.forename: Yusuke
authors.initials: Y
authors.lastname: Murakami
authors.forename: Takeshi
authors.initials: T
authors.lastname: Maeda
authors.forename: Yasushi
authors.initials: Y
authors.lastname: Shiomi
mesh_terms: D018929:Cell Culture Techniques / Q000379:methods; D002453:Cell Cycle; D018797:Cell Cycle Proteins / Q000302:isolation & purification* / Q000378:metabolism; D005591:Chemical Fractionation / Q000379:methods*; D002843:Chromatin / Q000302:isolation & purification* / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D064110:Minichromosome Maintenance Proteins / Q000302:isolation & purification / Q000378:metabolism; D011485:Protein Binding
chemical_list: D018797:Cell Cycle Proteins; D002843:Chromatin; D064110:Minichromosome Maintenance Proteins