Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.1.30.2 (endonuclease)
 18,621 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The genes for a Class II restriction-modification system (HhaII) from Haemophilus haemolyticus have been cloned in Escherichia coli. The vector used for cloning was plasmid pBR322 which confers resistance to tetracycline and ampicillin and contains a single endonuclease R-PstI site, (formula: see text), in the ampicillin gene. The procedure developed by Bolivar et al. (1977) was used to form DNA recombinants. H. haemolyticus DNA was cleaved with PstI endonuclease and poly(dC) extensions were added to the 3'-OH termini using terminal deoxynucleotidyl transferase. Circular pBR322 DNA was cleaved to linear molecules with PstI endonuclease and poly(dG) extensions were added to the 3'-OH termini, thus regenerating the PstI cleavage site sequences. Recombinant molecules, formed by annealing the two DNAs, were used to transfect a restriction and modification-deficient strain of E. coli (HB101 r-m-recA). Tetracycline-resistant clones were tested for acquisition of restriction phenotype (as measured by growth on plates seeded with phage lambdacI-0). A single phage-resistant clone was found. The recombinant plasmid, pD110, isolated from this clone, had acquired 3 kilobases of additional DNA which could be excised with PstI endonuclease. In addition to the restriction function, cells carrying the plasmid expressed the HhaII modification function. Both activities have been partially purified by single-stranded DNA-agarose chromatography. The cloned HhaII restriction activity yields cleavage patterns identical to HinfI. A restriction map of the cloned DNA segment is presented.
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PMID:Cloning of restriction and modification genes in E. coli: the HbaII system from Haemophilus haemolyticus. 35 Jul 14

A restriction endonuclease cleavage map of the tetracycline resistance plasmid pAB124, originally isolated from Bacillus stearothermophilus, was constructed using ten enzymes. Tetracycline resistance was associated with a 1 x 95 megadalton (Md) region of pAB124 lying between two EcoRI sites, and this region was circularized to produce a viable tetracycline resistance plasmid (pAB224), with two EcoRI fragments of pAB124 deleted amounting to 0 x 95 Md. A second plasmid (pAB524) with one EcoRI fragment (0 x 6 Md) of pAB124 deleted was also constructed. Restriction endonuclease cleavage maps of pAB224 and pAB524 were constructed.
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PMID:Characterization of Bacillus stearothermophilus plasmid pab124 and construction of deletion variants. 625 Nov 58

Ten plasmids were isolated as covalently closed circular deoxyribonucleic acid from antibiotic-resistant thermophilic bacteria. Of the 10 plasmids tested, 2 could transform Bacillus subtilis, yielding resistance to specific antibiotics. Plasmid pTB20 (2.8 X 10(6) daltons, approximately 24 copies per chromosome) specifies resistance to tetracycline (Tcr), whereas pTB19 (17.2 X 10(6) daltons, approximately 1 copy per chromosome) renders the host resistant to both kanamycin and tetracycline (KMrTcr). Three plasmids were not self-transmissible. The restriction endonuclease cleavage maps of the two plasmids, pTB19 and pTB20, were constructed. pTB19 and pTB20, both of which were originally isolated from thermophilic bacilli, were tested for stability in B. subtilis. Digestion of pTB19 followed by ligation yielded deletion plasmids pTB512 (Kmr), pTB52 (Tcr), and pTB53 (KmrTcr). Determinants of Kmr, Tcr, and DNA replication were associated with EcoRI fragments R1b (4.2 X 10(6) daltons), R3 (2.8 X 10(6) daltons), and R1a (4.2 X 10(6) daltons), respectively. Restriction endonuclease cleavage maps of pTB51, pTB52, and pTB53 were constructed. Tetracycline resistance of pTB20 was confirmed to be in the EcoRI fragment (1.85 X 10(6) daltons).
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PMID:Isolation and characterization of antibiotic resistance plasmids from thermophilic bacilli and construction of deletion plasmids. 626 56

Tetracycline-resistance (TCr) plasmid pTP-5 (4.46 kb) from Staphylococcus aureus was cleaved with HindIII into three fragments: A (2.35 kb), B (1.57 kb) and C (0.54 kb). A deletion plasmid (3.92 kb) lacking HindIII-C fragment was obtained, and was designated pNS1. This plasmid retained the TCr phenotype and the ability to replicate autonomously in Bacillus subtilis. A restriction endonuclease cleavage map of pNS1 was constructed. Attempts to construct smaller plasmids by digesting pNS1 with BAL31 nuclease led to production of a set of deletion derivatives whose molecular sizes range from 3.75 to 2.77 kb. Through analyses of these derivatives, the regions essential for autonomous replication and expression of TCr were deduced.
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PMID:Construction and propagation of deletion derivatives of staphylococcal tetracycline-resistance plasmid pTP-5 in Bacillus subtilis. 630 40

A homozygous a2/a2 and b4/b4 rabbit has been hyperimmunized with Micrococcus lysodeikticus. Poly(A)-containing RNA has been isolated from the spleen and translated in vitro, and translation products have been analyzed by NaDodSO4/polyacrylamide gel electrophoresis. Double-stranded cDNA has been synthesized from poly(A)-containing RNA template and inserted in the Pst I endonuclease site of plasmid pBR322 by using the oligo(dC).oligo(dG) tailing procedure. Tetracycline-resistant ampicillin-sensitive clones containing cDNA complementary to a kappa light chain mRNA have been selected by differential screening and their ability to hybridize to a spleen mRNA having the same size as a mouse kappa light chain mRNA. Two clones, pRk-15 and pRk-32, have been selected to determine the nucleotide sequence of the constant and 3' untranslated regions of kappa light chain mRNA, by the Maxam and Gilbert partial degradation method. Comparison of homologous regions of mouse kappa chain mRNA and b4 rabbit kappa chain mRNA reveals 61% homology in the constant region and 59% homology in the 3' untranslated region.
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PMID:Nucleotide sequence of constant and 3' untranslated regions of a kappa immunoglobulin light chain mRNA of a homozygous b4 rabbit. 679 36

The transfer of tetracycline resistance among strains of Clostridium difficile is described. Transfer occurred by a conjugation-like event that was insensitive to deoxyribonuclease, could not be mediated by donor culture filtrates or chloroform-treated donor cultures, and required cell-to-cell contact. Tetracycline-resistant progeny recovered from matings displayed a resistance phenotype identical to that of the donor in level of resistance, constitutive expression, and transmissibility. Although the original tetracycline-resistant donor contained 5 x 10(6)- and 22 x 10(6)-dalton plasmids, standard physical analyses of antibiotic-resistant transconjugants revealed no plasmid deoxyribonucleic acid molecules in common with the donor strain. Furthermore, tetracycline-susceptible derivatives of the original donor always possessed a plasmid complement identical to that of the resistant parental strain as determined by restriction endonuclease digestion analysis. The results indicate that the tetracycline resistance determinant(s) was not encoded by readily detectable plasmid deoxyribonucleic acid and may be chromosomally located.
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PMID:Transferable tetracycline resistance in Clostridium difficile. 727 Dec 79

The conjugative transposon Tn916 was used for mutagenesis of Clostridium acetobutylicum ATCC 824. Tetracycline-resistant mutants were screened for loss of granulose synthesis and five classes of granulose mutants, that contained single transposon insertions, were identified on the basis of altered solvent production. Class 1 mutants did not make acetone or butanol, lacked activity of enzymes induced during solventogenesis, and did not sporulate, indicating that they are regulatory mutants. The class 2 mutant strains also did not produce acetone but did form small amounts of butanol and ethanol, while the class 3 mutants produced low amounts of all solvents. Class 4 and 5 mutants produced essentially the same or higher amounts of solvents than the parent strain. Transposon insertions in the class 1 mutants were used as markers for in vitro synthesis of flanking chromosomal DNA using Tn916-specific primers. The DNA fragments were labeled to produce specific probes. Transposon insertion sites in the chromosomes of 13 different class 1 regulatory mutants were compared by hybridization of the specific probes to Southern blots of restriction endonuclease-digested parental chromosomal DNA. Insertions in two mutants appeared to be in the same region of the chromosome. These results predict that multiple regulatory elements are required to induce solvent production and sporulation.
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PMID:Analysis of Tn916-induced mutants of Clostridium acetobutylicum altered in solventogenesis and sporulation. 776 50

Tetracycline-resistance in gram-negative periodontal bacteria is often due to the presence of the tet(Q) gene. In the present study the polymerase chain reaction (PCR) was used to examine 54 isolates of gram-negative anaerobic rods (Prevotella intermedia, Prevotella nigrescens and related or Bacteroides-like species) for the presence of the tet(Q) gene. The isolates were recovered from 42 patients with periodontal disease living in northern Europe and North America. An 814 base-pair segment of the tet(Q) gene was amplified from all 41 isolates resistant to tetracycline with minimal inhibitory concentrations of 4 micrograms/ml and above. The presence of the tet(Q) gene was verified using hybridization with a specific oligonucleotide internal to the amplified region and restriction endonuclease digestion with DdeI. A PCR product of the same size was also amplified from one tetracycline susceptible isolate (minimal inhibitory concentration = 0.5 microgram/ml). However, this isolate and the one isolate that was resistant to tetracycline at 4 micrograms/ml showed a weaker signal than the remaining isolates when hybridized with the internal probe. Typing of the PCR products using restriction endonuclease digests with AluI and HpaII revealed two clusters of distinct electrophoresis patterns, indicating that two different subtypes of the tet(Q) gene were present in this material. A control strain containing the tet(Q) gene from Bacteroides thetaiotaomicron had a different electrophoresis pattern for AluI. This study indicated that subtypes of the tet(Q) gene in tetracycline-resistant gram-negative periodontal bacteria exist both within the same patient and within the same species.
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PMID:Tetracycline resistance in Prevotella isolates from periodontally diseased patients is due to the tet(Q) gene. 902 55

Double-stranded cDNA has been synthesized from a polyadenylylated potato spindle tuber viroid (PSTV) template and inserted in the Pst I endonuclease site of plasmid pBR322 by using the oligo(dC).oligo(dG)-tailing procedure. Tetracycline-resistant ampicillin-sensitive transformants contained sequences complementary to PSTV [(32)P]cDNA, and one recombinant clone (pDC-29) contains a 460-base-pair insert. This cloned double-stranded PSTV cDNA contains the cleavage sites for six restriction endonucleases predicted by the published primary sequence of PSTV as well as one additional site each for Ava I, Hae III, Hpa II, and Sma I. The additional Ava I, Hpa II, and Sma I sites are explained by the presence of a second C-C-C-G-G-G sequence in the cloned double-stranded cDNA. The largest fragment released by Hae III digestion contains approximately 360 base pairs. These results suggest that we have cloned almost the entire sequence of PSTV, but the sequence cloned differs slightly from that published. Hybridization probes derived from pDC-29 insert have allowed detection and preliminary characterization of RNA molecules having the same size as PSTV but the opposite polarity. This RNA is present during PSTV replication in infected tomato cells.
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PMID:Molecular cloning and characterization of potato spindle tuber viroid cDNA sequences. 1659 77