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Zoekopdracht: faculteit: "FNWI" en publicatiejaar: "2005"

AuteursS.P. Elmore, M. Muller, N.O.E. Vischer, T. Odijk, C.L. Woldringh
TitelSingle-particle tracking of oriC-GFP fluorescent spots during chromosome segregation in Escherichia coli.
TijdschriftJournal of Structural Biology
FaculteitFaculteit der Natuurwetenschappen, Wiskunde en Informatica
Instituut/afd.FNWI: Swammerdam Institute for Life Sciences (SILS)
TrefwoordenDNA segregation; Escherichia cloi; Mean square displacement; Random and cinfined diffusion; Simulations
Basisclassificatie35.73 biochemistry: molecular genetics
SamenvattingDNA regions close to the origin of replication were visualized by the green fluorescent protein (GFP)-Lac repressor/lac operator system. The number of oriC-GFP fluorescent spots per cell and per nucleoid in batch-cultured cells corresponded to the theoretical DNA replication pattern. A similar pattern was observed in cells growing on microscope slides used for time-lapse experiments. The trajectories of 124 oriC-GFP spots were monitored by time-lapse microscopy of 31 cells at time intervals of 1, 2, and 3 min. Spot positions were determined along the short and long axis of cells. The lengthwise movement of spots was corrected for cell elongation. The step sizes of the spots showed a Gaussian distribution with a standard deviation of 110 nm. Plots of the mean square displacement versus time indicated a free diffusion regime for spot movement along the long axis of the cell, with a diffusion coefficient of 4.3 ? 2.6 ? 10?5 ?m2/s. Spot movement along the short axis showed confinement in a region of the diameter of the nucleoid (800 nm) with an effective diffusion coefficient of 2.9 ? 1.7 ? 10?5 ?m2/s. Confidence levels for the mean square displacement analysis were obtained from numerical simulations. We conclude from the analysis that within the experimental accuracy ? the limits of which are indicated and discussed ? there is no evidence that spot segregation requires any other mechanism than that of cell (length) growth.
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