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Biomedical subjects

W Goebel

Publications and source records attributed to W Goebel.

At least 253 records · Page 14Linked to original sources
PubMed1977

Transposition and insertion of intact, deleted and enlarged ampicillin transposon Tn3 from mini-R1 (Rsc) plasmids into transfer factors.

The miniR1-(Rsc)-plasmids which derive from the copy mutant R1drd-19B2 (pKN102) are non-conjugative extrachromosomal elements which can not be co-transferred by various transfer factors to recipient strains under standard mating conditions. The attempts to mobilize Rsc11 by F'lac lead to transconjugants carrying F'lac::Tn3 with Tn3 mainly inserted into the lac operon. In addition it can be shown that Rsc11 can become inserted as a complete unit into the transfer factor giving rise to rather unstable recombinant intermediates. Dissociation of these intermediates may lead to alterations of the original plasmids. The Tn3 part of Rsc13 can be enlarged or deleted by in vitro manipulations. In vitro insertion of EcoRI-fragments into an EcoRI+ site of Tn3 leads to new transposable units which can be transposed to the RTF part of R1. This new genetic entity can be stably integrated into the chromosome of E. coli by integrative suppression of a dnaAts-mutation. Deletions at one end or the central region of Tn3 abolish the capability of transposition. However, the Rsc-plasmids containing the deleted Tn3 can still be inserted into the transfer factor as complete units. The resulting recombinants are unstable leading after dissociation in some cases to new plasmids with altered properties.

Ampicillin↗
PubMed1977

Replication of the mini-R1 plasmid Rsc11 and Rsc11 hybrid plasmids.

Replication of the multicopy mini-R1 plasmid, Rsc11, is dependent on host replication functions dna A, B, C, E and G but independent of polA1. Chloramphenicol immediately stops its replication. A stable relaxation complex is not formed. Composite plasmids were constructed with Rsc11 and other small replicons like pSC101, ColE1 and mini-ColE1. In all combinations the amount of hybrid plasmid DNA in the cell never exceeds the amount of Rsc11 DNA itself. This leads to varying copy numbers of the hybrid plasmids depending on the size of the second plasmid. Replication of the composite plasmids proceeds probably always under the control of the Rsc11 part although the second replicon is still functional. The composite plasmids are incompatible with both the parent replicons.

Chloramphenicol↗
PubMed1977

Properties of hybrid plasmids, consisting of parts of the mini-R1 factor Rsc11 and ColE1.

In vitro joining of the two small multicopy plasmids Rsc11 and ColE1 by a poly dAdT linker resulted in hybrid plasmids, which determine resistance to ampicillin and immunity to colicin E1. Isolation of the plasmid DNA from single colonies revealed that a variety of hybrid plasmids was formed. Cleavage of these plasmids with restriction endonucleases HinII, HindIII, EcoRI, SmaI and BamI and hybridization with ColE1 demonstrated that they contain different parts of the parent plasmids, Rsc11 and ColE1. Their copy number in the cell is between 6 and 15 per chromosome depending on the plasmid. None of these plasmids can replicate in polA mutants. Replication continues in the presence of chloramphenicol. This suggests that replication can only occur from the ColE1 origin and that the replication function of the Rsc11 part is lost. The hybrid plasmids are compatible with Rsc11 but not with ColE1. The comparison of the physical maps of these Rsc11--ColE1 hybrids with their functions allows a partial determination of the location of ampicillin resistance, replication and incompatibility on the Rsc11 genome.

Ampicillin↗
PubMed1977

Escherichia coli membrane proteins with an affinity for deoxyribonucleic acid.

From the membrane fraction of Escherichia coli K-12 strain, four protein fractions (peaks I, IIa, IIb, and III) which have affinity for deoxyribonucleic acid (DNA) have been isolated. The molecular weights of these proteins are between 12,000 and 8,000. Only the peak III fraction contains a protein that binds preferentially to single-stranded DNA, whereas the others contain proteins that bind also to double-stranded DNA. The binding activity of the peak IIb protein is inhibited in the presence of polyuridylic acid. Peak I and peak IIa protein fractions behave like hydrophobic proteins.

Bacterial Proteins↗
PubMed1977

Characterization of a plasmid from Streptomyces coelicolor A3(2).

Covalently closed circular deoxyribonucleic acid (DNA) with a molecular weight of 20 X 10(6) was identified in strains of Streptomyces coelicolor A3(2) of various fertility types. Hybridization studies and digestion by various restriction endonucleases indicated that the circular DNAs (pSH1) were identical regardless of the fertility type (UF, IF, or NF) of the strain from which it was isolated. The pSH1 DNA was cleaved to many fragments by the endonucleases HincII, SmaI, and SalI and to three or four fragments by BamHI and PstI. Plasmid pSH1 carries single sites for each of the two restriction enzymes, EcoRI and HindIII. These sites are 7.6 X 10(6) daltons apart. Attempts to isolate the fertility factor SCP1 as covalently closed circular DNA were unsuccessful. These data suggest that the biochemically isolated plasmid pSH1 is not identical to the genetically characterized fertility factor SCP1, which has been identified in an autonomous state in IF-type strains and in an integrated state in NF-type strains.

Bacteriocins↗
PubMed1976

Cloning of calf thymus satellite I DNA in Escherichia coli.

The 1400 base pair repeat produced by digestion of calf satellite I DNA (phi = 1.714 g/cm3) with EcoRI, was cloned in E. coli. The hybrid plasmid (pGM 214) which contains the ColE1-Ap vector (pSF 2124) and the 1400 base pair fragment replicates stably in E. coli and can be amplified by chloramphenicol treatment. No clone was found in which more than one "repeat unit" of the satellite I DNA was present in the chimaera plasmid. Digestion of the original satellite I and the plasmid pGM 214 with R-SmaI shows that the satellite DNA replicated in E. coli is cleaved by the restriction endonuclease SmaI whereas the original satellite I DNA from calf thymus is not, suggesting that the satellite I contains a large amount of modified cytosine or guanosine, probably 5-methyl-cytosine. R-EcoRI* produces a number of fragments with the satellite I in the range of 300 base pairs to 1400 base pairs. A physical map of pGM 214 (and pSF 2124) with R-EcoRI, R-HincII, R-HindIII, R-SmaI, R-BamI and R-EclI was constructed. The 1400 base pair "repeat unit" in the pGM 214 is efficiently transcribed in vitro by purified RNA polymerase, starting from a pSF 2124 promoter. The restriction enzyme EclI produces a 350 base pair repeat with calf satellite II (phi = 1,722 g/cm3), whereas the satellite I is not cut by this enzyme.

Animals↗
PubMed1976

Plasmids controlling synthesis of hemolysin in Escherichia coli. II. Polynucleotide sequence relationship among hemolytic plasmids.

Plasmids of three different sizes, designated as plasmid A (mw: 65 X 10(6), plasmid B (mw: 41 X 10(6) and plasmid C (mw: 32 X 10(6) respectively, have been isolated from various hemolytic wild-type strains of E. coli. DNA-DNA hybridization was performed to determine their relationship. The wild-type strain, PM167a, harbours plasmids of all three sizes. Hybridization studies indicate that all three plasmids share extented sequence homologies but that plasmid A is not composed of plasmids B and C. Hybridization between plasmids of the donor strain and those of appropriate transconjugants demonstrates that in some cases plasmids with identical size are not longer completely homologous in their nucleotide sequences. This indicates that despite their defined sizes these plasmids are not stable genetic entities, but rather they undergo frequently recombination and dissociation during conjugation. In one particular transconjugant strain, K12-PM152/1, a plasmid D was found which is a stable recombined molecule of plasmids B and C of the original strain. Plasmids of size B found as the only extrachromosomal elements in a hemolytic wild-type strain (P224) and two transconjugant strains (e.g. K12-CM20 and K12-PM167/1) share extended nucleotide sequence homologies but are not identical. Little sequence homology was observed between two different hemolytic plasmids and the F and the Col Ib plasmids suggesting that the former do not belong to either the F-like or the I-like group of plasmids. Another hemolytic plasmid is F-like based on its sequence homologies with the F factor.

Base Sequence↗
PubMed1975

The fate of a bacterial plasmid in mammalian cells.

When hamster cells are infected with the bacterial plasmid colicinogenic factor E1 (ColE1), as much as 5-8% of the input plasmid radioactivity is found in the recipient cell, mainly in the nuclear fraction. Density shift experiments with bromodeoxyuridine labeled ColE1 DNA indicate that part of the input DNA may be replicated in the nucleus. ColE1 specific RNA but no colicin E1, can be detected during the first two generations after the uptake of ColE1 DNA. However, extrachromosomal ColE1 DNA is unstable in the mammalian cells and is degraded to acid soluble fragments after a few generations.

Animals↗
PubMed1975

Class of small multicopy plasmids originating from the mutant antibiotic resistance factor R1 drd-19B2.

The large mutant R-factor R1drd-19B2 gives rise to several classes of small, covalently closed circular deoxyribonucleic acids (DNAs), designated as Rsc DNAs, when harbored by the K-12 strain CRT46 which carries a dnaA mutation. The molecular weights of these DNA molecules range from 3 X 106 to 8.4 X 106. Cells arising from single colonies of CRT46-R1drd-19B2 harbor only one to two copies of the large mutant R-factor and in addition 10 to 20 copies of Rsc plasmid of a discrete size class per chromosome. The larger Rsc DNAs carry the ampicillin resistance gene. After transformation the small circular DNAs are present in Escherichia coli C in a large number of copies, up to 100 copies per chromosome. Hybridization studies between Rsc plasmids indicate that they possess common DNA sequences.

Ampicillin↗