Abstract:Five criteria of genetic relatedness are considered. The first, transfer of plasmids between groups, is frequently not a good criterion, because transfer is possible between all genera of the Enterobacteriaceae and also to genera in other families. Though transfer to closely related groups is most frequent, host restriction and the properties of the plasmid may influence the transfer frequency as much as the relatedness of the donor and recipient. The second criterion is interspecies recombination (integration… Show more
“…3B), confirming the position of the 4.1-kb band BlnI-F near rmD. DISCUSSION For many years there has been a recognition that the chromosomal structure and gene order in the linkage maps of S. typhimurium and E. coli K-12 are strongly conserved (19,37,38,44); we have now confirmed this conservation by analysis of genomic cleavage maps of these two organisms. A genomic cleavage map of E. coli K-12 was developed initially for NMtI (52), and since then similar maps have been developed for AvrII (=BlnI) (11,36), NotI (14), SfiI (35), XbaI (36), and CeuI (26); much but not all of the latter work has been with the K-12 strain MG1655.…”
Endonuclease digestion of the 4,800-kb chromosome of SabloneUa lyphimurium LT2 yielded 24 XbaI fragments, 12 BIn! fragments, and 7 CeuI fragments, which were separated by pulsed-field gel electrophoresis.The 90-kb plasmid pSLT has oneXbaI site and one BinI site. The locations of the fragments around the circular chromosome and of the digestion sites of the different endonucleases with respect to each other were determined by excision of agarose blocks containing fragments from single digestion, redigestion with a second enzyme, end labelling with 32P by using T7 DNA polymerase, reelectrophoresis, and autoradiography. Forty-three cleavage sites were established on the chromosome, and the fragments and cleavage sites were designated in alphabetical order and numerical order, respectively, around the chromosome. One hundred nine independent TnlO insertions previously mapped by genetic means were located by pulsed-field gel electrophoresis on the basis of the presence of XbaI and Bin1 sites in TnlO. The genomic cleavage map was divided into 100 units called centisomes; the endonuclease cleavage sites and the genes defined by the positions of Tn1O insertions were located by centisome around the map. There is very good agreement between the genomic cleavage map, defined in centisomes, and the linkage map, defined in minutes. All seven rRNA genes were located on the map; all have the Ceu! digestion site, all four with the tRNA gene for glutamate have theXbaI and the BinI sites, but only four of the seven have the Bin! site in the 16S rRNA (rrs) gene. Their inferred orientation of transcription is the same as in Escherichia coli. A rearrangement of the rrnB and rrnD genes with respect to the arrangement in E. coli, observed earlier by others, has been confirmed. The sites for all three enzymes in the rrn genes are strongly conserved compared with those in E. coli, but the XbaI and BinI sites outside the mrn genes show very little conservation.
“…3B), confirming the position of the 4.1-kb band BlnI-F near rmD. DISCUSSION For many years there has been a recognition that the chromosomal structure and gene order in the linkage maps of S. typhimurium and E. coli K-12 are strongly conserved (19,37,38,44); we have now confirmed this conservation by analysis of genomic cleavage maps of these two organisms. A genomic cleavage map of E. coli K-12 was developed initially for NMtI (52), and since then similar maps have been developed for AvrII (=BlnI) (11,36), NotI (14), SfiI (35), XbaI (36), and CeuI (26); much but not all of the latter work has been with the K-12 strain MG1655.…”
Endonuclease digestion of the 4,800-kb chromosome of SabloneUa lyphimurium LT2 yielded 24 XbaI fragments, 12 BIn! fragments, and 7 CeuI fragments, which were separated by pulsed-field gel electrophoresis.The 90-kb plasmid pSLT has oneXbaI site and one BinI site. The locations of the fragments around the circular chromosome and of the digestion sites of the different endonucleases with respect to each other were determined by excision of agarose blocks containing fragments from single digestion, redigestion with a second enzyme, end labelling with 32P by using T7 DNA polymerase, reelectrophoresis, and autoradiography. Forty-three cleavage sites were established on the chromosome, and the fragments and cleavage sites were designated in alphabetical order and numerical order, respectively, around the chromosome. One hundred nine independent TnlO insertions previously mapped by genetic means were located by pulsed-field gel electrophoresis on the basis of the presence of XbaI and Bin1 sites in TnlO. The genomic cleavage map was divided into 100 units called centisomes; the endonuclease cleavage sites and the genes defined by the positions of Tn1O insertions were located by centisome around the map. There is very good agreement between the genomic cleavage map, defined in centisomes, and the linkage map, defined in minutes. All seven rRNA genes were located on the map; all have the Ceu! digestion site, all four with the tRNA gene for glutamate have theXbaI and the BinI sites, but only four of the seven have the Bin! site in the 16S rRNA (rrs) gene. Their inferred orientation of transcription is the same as in Escherichia coli. A rearrangement of the rrnB and rrnD genes with respect to the arrangement in E. coli, observed earlier by others, has been confirmed. The sites for all three enzymes in the rrn genes are strongly conserved compared with those in E. coli, but the XbaI and BinI sites outside the mrn genes show very little conservation.
“…Early versions of the S. typhimurium linkage map were 138 min of entry time (29), but for convenience of comparison with E. coli, later editions have used 100 min (31) (sometimes called "units" to emphasize that these are not really minutes of Hfr entry time). The order of genes on the linkage maps of E. coli and S. typhimurium has long been known to be very similar (28). The (13,23,24).…”
XbaI digestion and pulsed-field gel electrophoresis of the genome of SalmoneUla typhimurium LT2 yields 24 fragments: 23 fragments (total size, 4,807 kb) are from the chromosome, and one fragment (90 kb) is from the virulence plasmid pSLT. Some of the 23 fragments from the chromosome were located on the linkage map by the use of cloned genes as probes and by analysis of strains which gain an XbaI site from the insertion of TnlO. Twenty-one of the fragments were arranged as a circular physical map by the use of linking probes from a set of 41 lysogens in which Mud-P22 was stably inserted at different sites of the chromosome; fragment W (6.6 kb) and fragment X (6.4 kb) were not located on the physical map. XbaI digestion of strains with TnlO insertions allowed the physical locations of specific genes along the chromosome to be determined on the basis of analysis of new-fragment sizes. There is good agreement between the order of genes on the linkage map, which is based primarily on P22 joint transduction and F-mediated conjugation, and the physical map, but there are frequently differences in the length of the interval from the two methods. These analyses allowed the measurement of the amount of DNA packaged in phage P22 heads by Mud-P22 lysogens following induction; this varies from ca. 100 kb (2 min) to 240 kb (5 min) in different parts of the chromosome.
“…In view of the considerable time-length of chromosome already transferred, it appears inevitable that the statement about the map order of P . mirabilis differing markedly from that of E. coli (see Sanderson, 1976) will stand. Marked differences are in accord with the fact that Brenner et al (1969) only detected 6 yo relatedness between E. coli DNA and that of P .…”
Section: Proteus Chromosome Transfer 161mentioning
Mating procedures for detection of mobilization of the Proteus mirabilis chromosome were re-investigated. The chromosome was mobilized by plasmid D, the previously used hybrid between plasmids P-lac and Rldrdl9. About a 40-fold increase in recombinant recovery correlated with the absence of swarming during mating and a lower temperature of incubation. The modification introduced was that conjugation was allowed to proceed on a non-selective supplemented minimal medium at 30 "C before washing and plating on selective media. Final incubation was also at 30 "C. This technique enabled eight additional chromosomal markers to be mapped. Polarized transfer of the chromosome was shown by gradient of transmission experiments using a previously described marker as reference, by linkage analysis with reference to proximal and distal markers and (less successfully) by interrupted mating on solid medium. Markers of plasmid D transferred at high frequency to all recombinants. The plasmid was stable in recombinants and could transfer itself and chromosomal markers of the new hosts in further matings. Resulting recombination of markers occurred at usual frequencies. The marker order, his-1, pyrBl, cysAl, cysGl, cysC1, argA2, metF2, nalA1, leuB2, did not resemble the order of these markers in Escherichia coli.
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