Replication initiates in a broad zone in the amplified CHO dihydrofolate reductase domain

• Published in: Cell Vol. 61; no. 6; pp. 1075 - 1087
• Main Authors: Vaughn, James P., Dijkwel, Pieter A., Hamlin, Joyce L.
• Format: Journal Article
• Language: English
• Published: Cambridge, MA Elsevier Inc 15.06.1990; Cell Press
• Subjects: Animals; Biological and medical sciences; Cell Line; Cricetinae; Cricetulus; DNA - genetics; DNA - isolation & purification; DNA Replication; Electrophoresis, Gel, Two-Dimensional; Female; Fundamental and applied biological sciences. Psychology; Gene Amplification; Molecular and cellular biology; Molecular genetics; Ovary; Replication; Restriction Mapping; Tetrahydrofolate Dehydrogenase - genetics; Transfection; Site specificity; Cell culture; Gel electrophoresis; Enzyme; Replication initiation; Enzymatic digestion; Tetrahydrofolate dehydrogenase; Localization; CHO cell line; Ion exchange chromatography
• ISSN: 0092-8674; 1097-4172
• DOI: 10.1016/0092-8674(90)90071-L

We have used two complementary two-dimensional gel electrophoretic methods to localize replication initiation sites and to determine replication fork direction in the amplified 240 kb dihydrofolate reductase domain of the methotrexate-resistant CHO cell line CHOC 400. Surprisingly, our analysis indicates that replication begins at many sites in several restriction fragments distributed throughout a previously defined 28 kb initiation locus, including a fragment containing a matrix attachment region. Initiation sites were not detected in regions lying upstream or downstream of this locus. Our results suggest that initiation reactions in mammalian chromosomal origins may be more complex than in the origins of simple microorganisms.