Relationships between the physical and genetic maps of a 470 × 103 base-pair region around the terminus of Escherichia coli K12 DNA replication

Bouché, Jean‐Pierre; Gélugne, Jean‐Paul; Louarn, Jean-Michel; Kaiser, Kim

doi:10.1016/0022-2836(82)90414-4

Cited by 75 publications

(54 citation statements)

References 24 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…recE mutations reduce elevated levels of conjugational and plasmid recombination and UV and mitomycin C resistance produced by sbcA mutations in a recB recC genetic background (8,10,11,14,19,26,31,32). This supports the hypothesis that exoVIII plays a role in the processes of recombination and repair in recB recC sbcA mutants (3).The relationship of sbcA to recE has previously been investigated by Southern blot analysis of restriction endonuclease-digested chromosomal DNA (23,24 (5,9,13,15,23,29,35). The Southern blot analysis also revealed that mutations called sbcA differ; point mutations and one large deletion were found.…”

supporting

confidence: 76%

Analysis of the recE locus of Escherichia coli K-12 by use of polyclonal antibodies to exonuclease VIII

1988

View full text Add to dashboard Cite

Exonuclease VIII (exoVIII) of Escherichia coli has been purified from a strain carrying a plasmid-encoded recE gene by using a new procedure. This procedure yielded 30 times more protein per gram of cells, and the protein had a twofold higher specific activity than the enzyme purified by the previously published procedure (J. W. Joseph and R. Kolodner, J. Biol. Chem. 258:10411-10417, 1983). The sequence of the 12 N-terminal amino acids was also obtained and found to correspond to one of the open reading frames predicted from the nucleic acid sequence of the recE region of Rac (C. Chu, A. Templin, and A. J. Clark, manuscript in preparation). Polyclonal antibodies directed against purified exoVIII were also prepared. Cell-free extracts prepared from strains containing a wide range of chromosomal-or plasmid-encoded point, insertion, and deletion mutations which result in expression of exoVIII were examined by Western blot (immunoblot) analysis. This analysis showed that two point sbcA mutations (sbcA5 and sbcA23) and the sbc insertion mutations led to the synthesis of the 140-kilodalton (kDa) polypeptide of wild-type exoVIII. Plasmid-encoded partial deletion mutations of recE reduced the size of the cross-reacting protein(s) in direct proportion to the size of the deletion, even though exonuclease activity was still present. The analysis suggests that 39 kDa of the 140-kDa exoVIII subunit is all that is essential for exonuclease activity. One of the truncated but functional exonucleases (the pRAC3 exonuclease) has been purified and confirmed to be a 41-kDa polypeptide. The first 18 amino acids from the N terminus of the 41-kDa pRAC3 exonuclease were sequenced and found to correspond to one of the translational start signals predicted from the nucleotide sequence of racC (Chu et al., in preparation).sbcA mutations suppress the conjugation-mediated recombination deficiency, UV sensitivity, and mitomycin C sensitivity of Escherichia coli strains containing recB or recC mutations or both (3). In addition, an sbcA mutation in a recB recC mutant background stimulates the recombination of circular monomer and dimer and linear dimer plasmid substrates 20-to 100-fold and increases the efficiency of recombination-mediated transformation of linear dimer plasmids at least 100-fold (10,19,26,31).One additional phenotype associated with the presence of an sbcA mutation is the presence of an ATP-independent DNase, exonuclease VIII (exoVIII;3,25). ExoVIII is a highly processive 5'-to-3' exonuclease with a subunit molecular weight of 140,000 (21, 25). The enzyme preferentially degrades double-stranded linear DNA and has no activity on single-stranded DNA or double-stranded circular DNA in a supercoiled, nicked, or gapped configuration (21). ExoVIII is functionally similar to A exonuclease (6,21,22,25,27), and can substitute for the X red functions (12). The structural gene for exoVIII has been termed recE (25). recE mutations reduce elevated levels of conjugational and plasmid recombination and UV and mitomycin C resistance produ...

show abstract

supporting

confidence: 76%

Analysis of the recE locus of Escherichia coli K-12 by use of polyclonal antibodies to exonuclease VIII

1988

View full text Add to dashboard Cite

show abstract

“…This places hipA in the recently bridged transduction gap (4,11) of the chromosome in which only a few functions have been mapped (5,6). The hipA locus is remote from other loci affecting murein metabolism (2).…”

mentioning

confidence: 99%

Molecular cloning and expression of hipA, a gene of Escherichia coli K-12 that affects frequency of persistence after inhibition of murein synthesis

Moyed¹,

Broderick²

1986

J Bacteriol

241

302

View full text Add to dashboard Cite

The hipA gene at 33.8 min on the Escherichia coli chromosome controls the frequency of persistence upon inhibition of murein synthesis; for strains bearing hipA+ the frequency is 10-6, and for hipA-strains the frequency is 10-2. hip' has been cloned by selection for a kanamycin resistance determinant at 33.9 min. hipA+ is dominant over hipA-in both recA+ and recA backgrounds. The smallest DNA insert which contains hipA+, as determined by the ability of the plasmids to complement hipA-strains, is 1,885 base pairs. Both orientations of hipA+ are obtained when the cloning site of vector is remote from strong promoters; both orientations complement hipA-, and both encode a unique peptide of 50,000 Mr. The probable direction of transcription has been deduced from the pattern of peptides encoded by plasmids from which either end of the insert and adjacent vector sequences have been deleted. This information and the recovery of only one orientation of hipA+ when the cloning site is close to a strong promoter suggest that a high level of expression of the gene is not tolerated by E. col.

show abstract

“…For regions spanning up to 50 kilobases (kb), restriction maps are routinely constructed and have become indispensable tools both for organizing existing data and for planning experiments. In a few favorable cases, maps as long as 600 kb have been constructed at a resolution of a few kb (1)(2)(3), but it has proven difficult to extend existing mapping methods beyond this range. Given that the average human chromosome contains 105 kb of DNA, there is a gross disparity between the present limits of restriction-mapping techniques and the sequence complexity of cellular genomes.…”

mentioning

confidence: 99%

Random-clone strategy for genomic restriction mapping in yeast.

Olson¹,

Dutchik²,

Graham³

et al. 1986

Proc. Natl. Acad. Sci. U.S.A.

362

192

View full text Add to dashboard Cite

An approach to global restriction mapping is described that is applicable to any complex source DNA. By analyzing a single restriction digest for each member of a redundant set of A clones, a data base is constructed that contains fragment-size lists for all the clones. The clones are then grouped into subsets, each member of which is related to at least one other member by a significant overlap. Finally, a tree-searching algorithm seeks restriction maps that are consistent with the fragment-size lists for all the clones in each subset. The feasibility of the approach has been demonstrated by collecting data on 5000 A clones containing random 15-kilobase inserts of yeast DNA. It is shown that these data can be analyzed to produce regional maps of the yeast genome, extending in some cases for over 100 kilobases. In combination with hybridization probes to previously cloned genes, these local maps are already useful for defining the physical arrangement of closely linked genes. They may in the future serve as building blocks for the construction of a continuous global map.Fifteen years after the discovery of site-specific restriction endonucleases, the restriction map has become the standard medium for displaying the functional organization of short segments of DNA. For regions spanning up to 50 kilobases (kb), restriction maps are routinely constructed and have become indispensable tools both for organizing existing data and for planning experiments. In a few favorable cases, maps as long as 600 kb have been constructed at a resolution of a few kb (1-3), but it has proven difficult to extend existing mapping methods beyond this range. Given that the average human chromosome contains 105 kb of DNA, there is a gross disparity between the present limits of restriction-mapping techniques and the sequence complexity of cellular genomes.In this paper, we describe a project that is aimed at constructing a restriction map of total nuclear DNA from the yeast Saccharomyces, whose haploid genome size has been estimated to be 1.5 x 104 kb (4). The data collection has involved picking X clones at random and measuring the sizes of the restriction fragments generated by a single digest of each clone. Because enough clones have been analyzed to provide redundant sampling of most regions of the genome, it is possible to extract mapping information from the unordered fragment-size lists by an algorithm that systematically imposes the requirement that the set of restriction fragments generated from a single clone must be contiguous.At the present stage of the yeast project, we have analyzed 5000 X clones with average insert sizes of 15 kb, thereby achieving a sampling redundancy on the order of five. Using the data, it is now possible to construct maps of from 20 to >100 kb around most arbitrary starting points in the yeast genome. These local maps have already defined the physical arrangement of clusters of genes that previously had only been linked genetically, and they point the way toward the development of a global phy...

show abstract

Relationships between the physical and genetic maps of a 470 × 103 base-pair region around the terminus of Escherichia coli K12 DNA replication

Cited by 75 publications

References 24 publications

Analysis of the recE locus of Escherichia coli K-12 by use of polyclonal antibodies to exonuclease VIII

Analysis of the recE locus of Escherichia coli K-12 by use of polyclonal antibodies to exonuclease VIII

Molecular cloning and expression of hipA, a gene of Escherichia coli K-12 that affects frequency of persistence after inhibition of murein synthesis

Random-clone strategy for genomic restriction mapping in yeast.

Contact Info

Product

Resources

About