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The deoxyribonucleic acid labeled by a procedure described previously for labeling the chromosomal terminus of B. subtilis 168 was substantially enriched for sequences homologous to bacteriophages SP beta and phi 3T, which integrate in the terminal region.
By making use of the sporulation process, the terminus region of the Bacillus subtilis chromosome has been labeled with [3H]thymine in a highly specific manner. The result achieved supports the view that B. subtilis spores contain only completed chromosomes.
The nucleoids of Escherichia coli and the spirochetes Borrelia burgdorferi and Borrelia hermsii, agents of Lyme disease and relapsing fever, were examined by epifluorescence microscopy of bacterial cells embedded in agarose and lysed in situ with detergent and protease. The typical E. coli nucleoid was a rosette in which 20 to 50 long loops of DNA emanated from a dense node of DNA. The percentages of cells in a population having nucleoids with zero, one, two, and three nodes varied with growth rate and growth phase. The borrelia nucleoid, in contrast, was a loose network of DNA strands devoid of nodes. This nucleoid structure difference correlates with the unusual genome of Borrelia species, which consists primarily of linear replicons, including a 950-kb linear chromosome and linear plasmids. This method provides a simple, direct means to analyze the structure of the bacterial nucleoid.
The RuvC protein is important for DNA recombination and repair in Escherichia coli. The present work shows that a ruvC null mutation introduced into a recBC sbcBC background causes severe defects in chromosome segregation and cell division. Both defects were found to result from abortive recombination initiated by the RecA protein.
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A short DNA segment located in the kanamycin resistance region of plasmid R1 promotes site-specific recombination and plasmid maintenance. This segment has been reduced to 100 bp and subsequently to 44 bp without losing these properties. It can recombine with a similar segment located in the terminus region of the Escherichia coli chromosome. It is proposed that this recombination is responsible for the plasmid maintenance properties of the R1 segment. The chromosomal site has been isolated; it also shows site-specific recombination activity. Sequence homologies were also found with a phage site-specific integration locus in the chromosome of Xanthomonas campestris and with the plasmid ColE1 site-specific recombination locus. The recombinase required in all these systems is probably XerC, an E. coli enzyme acting on the cer site of plasmid ColE1 for the conversion of plasmid dimers to monomers. It is postulated that site-specific recombination in the terminus region of the chromosome intervenes in the partitioning of the two daughter chromosomes.
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Two strains of Str. antibioticus producing oleandomycin were studied. Strain 326 was obtained from the initial laboratory strain and strain 1607 from strain 326 as a result of multistage selection aimed at increasing the antibiotic production capacity. Extrachromosomal ring DNA could be identified in strain 1607. The identified DNA was designated as eSA1 or extrachromosomal element of Streptomyces antibioticus 1. The molecular weight of this DNA is 21.3 Md. It is represented by 1 copy per chromosome. No eSA1 was isolated from strain 326 at CsCl-EtBr gradient. Hybridization studies according to Southern with the use as a probe of 32-eSA1 DNA showed that strain 326 contained 1 copy of eSA1 per chromosome in the integrated state. The hybridization studies, electron microscopy and analysis of the total DNA in CsC1-EtBr gradient showed that eSA1 in strain 1607 was tandemly multireplicated in the chromosome content. In the autonomous state its number was approximately equal to 1 copy per chromosome. The presence of eSA1 in strain 1607 in the autonomous state probably results from its segregation during homologous recombination due to tandem multireplication. The data are indicative of multiplication in strain 1607 of the chromosome fragment 23.3 Md in size. It is suggested that an increase in the oleandomycin production capacity of strain 1607 is associated with multiplication of the DNA fragment (eSA1) containing the genes determining production of the antibiotic.
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