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Cell-Cycle-Regulated Expression and Subcellular Localization of the Caulobacter crescentus SMC Chromosome Structural Protein. Rasmus B. Jensen, 2003.Structural maintenance of chromosomes proteins (SMCs) bind to DNA and function to ensure proper chromosome organization in both eukaryotes and bacteria . Caulobacter crescentus possesses a single SMC homolog that plays a role in organizing and segregating daughter chromosomes . Approximately 1,500 to 2,000 SMC molecules are present per cell during active growth, corresponding to one SMC complex per 6,000 to 8,000 bp of chromosomal DNA . Although transcription from the smc promoter is induced during early S phase, a cell cycle transcription pattern previously observed with multiple DNA replication and repair genes, the SMC protein is present throughout the entire cell cycle . Examination of the intracellular location of SMC showed that in swarmer cells, which do not replicate DNA, the protein forms two or three foci . Stalked cells, which are actively engaged in DNA replication, have three or four SMC foci per cell . The SMC foci appear randomly distributed in the cell . Many predivisional cells have bright polar SMC foci, which are lost upon cell division . Thus, chromosome compaction likely involves dynamic aggregates of SMC bound to DNA . The aggregation pattern changes as a function of the cell cycle both during and upon completion of chromosome replication . Application of New Primer-Enzyme Combinations to Terminal Restriction Fragment Length Polymorphism Profiling of Bacterial Populations in Human Feces. Koji Nagashima, 2003.New primer-enzyme combinations for terminal restriction fragment length polymorphism (T-RFLP) targeting of the 16S rRNA gene were constructed by using the T-RFLP analysis program (designated TAP T-RFLP) located at the Ribosomal Database Project website, and their performance was examined empirically . By using the fluorescently labeled 516f primer (Escherichia coli positions 516 to 532) and 1510r primer (positions 1510 to 1492), the 16S rRNA gene was amplified from human fecal DNA . The resulting amplified product was digested with RsaI plus BfaI or with BslI . When the T-RFLP was carried out with fecal DNAs from eight individuals, eight predominant operational taxonomic units (OTUs) were detected with RsaI and BfaI digestion and 14 predominant OTUs were detected with BslI digestion . The distribution of the OTUs was consistent with the results of the computer simulations with TAP T-RFLP . The T-RFLP analyses of the fecal DNAs from individuals gave characteristic profiles, while the variability of the T-RFLP profiles between duplicate DNA preparations from the same samples were minimal . This new T-RFLP method made it easy to predict what kind of intestinal bacterial group corresponded to each OTU on the basis of the terminal restriction fragment length compared with the conventional T-RFLP and, moreover, made it possible to identify the bacterial species that an OTU represents by cloning and sequencing .
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