RESUMO
The average cellular positions of the ftsQAZ region (2 min) and the minB region (26.5 min) during the cell cycle was determined by fluorescent in situ hybridization using the position of oriC as a reference point. At the steady-state growth conditions used, newborn cells had replicated about 50% of the chromosome. By measuring the distances of the labelled oriCs with respect to mid-cell, we found two well-separated average oriC positions in cells of newborn length. These average oriC positions moved further apart along with cell elongation. The cellular position of the ftsQAZ gene region resembled the position of oriC, although its average position was closer to mid-cell. In contrast, a single minB focus was observed at cell birth. Separated minB foci appeared towards the end of DNA replication. The average positions of oriC, ftsQAZ and minB relative to each other fitted a model in which DNA replication takes place in the cell centre and subsequent gene regions pass sequentially through this centre. We have interpreted the polarized orientation of the studied gene regions as a consequence of the mode of DNA segregation.
Assuntos
Proteínas de Bactérias/genética , Ciclo Celular/genética , Cromossomos Bacterianos/fisiologia , Proteínas do Citoesqueleto , Replicação do DNA , Proteínas de Escherichia coli , Escherichia coli/genética , Adenosina Trifosfatases/genética , Adenosina Trifosfatases/metabolismo , Proteínas de Bactérias/metabolismo , Ciclo Celular/fisiologia , Proteínas de Ciclo Celular , Núcleo Celular/fisiologia , Escherichia coli/fisiologia , Hibridização in Situ Fluorescente , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Origem de Replicação/genéticaRESUMO
The origin of replication of Escherichia coli, oriC, has been labeled by fluorescent in situ hybridization (FISH). The E. coli K12 strain was grown under steady state conditions with a doubling time of 79 min at 28 degrees C. Under these growth conditions DNA replication starts in the previous cell cycle at -33 min. At birth cells possess two origins which are visible as two separated foci in fully labeled cells. The number of foci increased with cell length. The distance of foci from the nearest cell pole has been measured in various length classes. The data suggest: i) that the two most outwardly located foci keep a constant distance to the cell pole and they therefore move apart gradually in line with cell elongation; and ii) that at the initiation of DNA replication the labeled origins occur near the center of prospective daughter cells.
Assuntos
Ciclo Celular/genética , Escherichia coli/citologia , Escherichia coli/genética , Origem de Replicação , Divisão Celular , Polaridade Celular , Replicação do DNA , Escherichia coli/metabolismo , Hibridização in Situ FluorescenteRESUMO
In an earlier fluorescent in-situ hybridization (FISH) study on petunia (ten Hoopen et al. 1996), we found a considerable discrepancy between the genetic map and the physical map with respect to T-DNA insertions on metaphase chromosomes. For some transgenes we found a preference to integrate near the telomeres. Here, we studied the spatial position of transgenes in interphase nuclei by FISH and 3D-confocal microscopy to elucidate a possible structural preference for the nuclear localization of transgenes. Three transgenes located near telomeres on three different metaphase chromosomes showed a much more internal distribution in interphase root meristem than the telomeres, whereas a proximal transgene appeared to be distributed in a random fashion. The results point to local differences in chromatin compacting along a chromosome. These differences might explain a preference for T-DNA insertion in distal regions of the chromosome.
