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)

Upon a 50% isopropanol treatment of phage fl in a 1 M NaCl solution protein A (gene 3 product)--DNA complex is precipitated while protein B (gene 8 product) was still solubilized. After such a treatment the DNA--protein complex containing 10--40% of protein A and less than 0.0025% of protein B was obtained. Evidence was obtained that there was no non-specific rearrangement of protein A during the isolation procedure. The complex was treated with endonuclease R.HAC III, followed by electrophoresis of the resulted fragments and estimation of the [14C] protein A (labeled with [14C]histidine) throughout the gel. The maximal radioactivity coincided with the DNA bands, being proportional to the DNA content in the respective bands. The data obtained indicate that protein A is iniformly arranged along the DNA molecule.
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PMID:[Complex of single-stranded phage DNA and protein--a product of bacteriophage fl gene 3. Distribution of gene 3 protein in the DNA molecule]. 66 19

Chlamydia psittaci is a diverse group of organisms that affects birds and mammals. The number of biovars is unknown, and less is known about the number of serovars. Our restriction endonuclease analysis indicates that there are at least 5 biovars including avian, abortion-enteritis, IPA, M56, and GPIC. Monoclonal antibody studies revealed 4 serovars in the avian biovar. Monoclonal antibody studies have not yet been performed to identify multiple serovars in the other biovars; however, microimmunoassay studies indicate that a number of serovars may exist in the abortion and arthritis biovars. Of the 4 avian serovars, 2 are of major importance in the US avian population. These 2 serovars, psittacine and turkey, are each associated with important host preferences and disease characteristics. The turkey isolates have all been associated with either a serious disease in birds or human beings or with major epizootics in turkeys, often resulting in human disease. The psittacine serovar has been associated with serious disease in human beings; however, human involvement is usually limited to sporadic cases following exposure to companion birds or pigeons. The other 2 serovars, German duck and WC, are single isolates and their distributions and disease characteristics are not known.
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PMID:Genetic, immunologic, and pathologic characterization of avian chlamydial strains. 268 4

Photoalkylation, the ultraviolet irradiation of DNA with isopropanol and di-tert-butylperoxide, causes a variety of base alterations. These include 8-(2-hydroxy-2-propyl)guanines, 8-(2-hydroxy-2-propyl)adenines and thymine dimers. An E. coli endonuclease against photoalkylated DNA was assayed by conversion of superhelical PM2 phage DNA to the nicked form. Enzyme activities were compared between extracts of strain BW9109 (xth-), lacking exonuclease III activity, and strain BW434 (xth-,nth-), deficient in both exonuclease III and endonuclease III. The endonuclease level in the double mutant against substrate photoalkylated DNA was under 20% of the activity in the mutant lacking only exonuclease III. Irradiation of the DNA substrate in the absence of isopropanol did not affect the activity in either strain. Analysis by polyacrylamide gel electrophoresis identified the sites of DNA cleavage by purified E. coli endonuclease III as cytosines, both in DNA irradiated at biologically significant wavelengths and in photoalkylated DNA. Neither 8-(2-hydroxy-2-propyl)purines, pyrimidine dimers, uracils nor 6-4'-(pyrimidin-2'-one)pyrimidines were substrates for the enzyme.
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PMID:Photoalkylated DNA and ultraviolet-irradiated DNA are incised at cytosines by endonuclease III. 352 39

A new procedure is developed for isolation of highly purified preparations of restrictional endonoucleases Bam HI and Eco RI by means of fractionation with isopropyl alcohol. Restrictional endonuclease Bam HI, practically free of unspecific nucleases, was isolated after ultrasonic destruction of cells, precipitation of the restrictases with isopropanol and chromatography on DEAE cellulose. Additional chromatography on hydroxyapatite enabled to obtain the homogenous preparation of Bam HI restrictase, as shown by polyacrylamide gel disc electrophoresis. Other organic solvents (acetone, ethanol) might be also used for purification of the restrictional endonucleases.
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PMID:[New means of isolating restriction endonuclease preparations using organic solvents]. 625 63

Based on the results of a systematic study of factors affecting plasmid yield and purity, a procedure suitable for the rapid screening for and isolation of covalently closed circular DNA from Streptomyces lividans and Escherichia coli was developed. The method consists of lysis of lysozyme-treated bacteria combined with alkaline denaturation of DNA at high temperature. Renaturation of CCC DNA and precipitation of single-stranded DNA together with protein is achieved by the addition of a minimal amount of phenol/chloroform. The screening procedure uses only a single tube and the samples can be analyzed by agarose gel electrophoresis about 30 min after lysis. Removal of phenol and further purification of the plasmid preparation is achieved by consecutive precipitations with isopropanol and spermine, followed by extraction with ethanol, producing samples suitable for restriction endonuclease digestion, ligation, and transformation of S. lividans protoplasts or competent E. coli cells in about 2 h. All steps of the procedure are explained in detail with information about the effects of changing parameters. This should help the experimenter to obtain reproducible results and may be useful if the method has to be adapted to new strains or plasmids.
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PMID:Factors affecting the isolation of CCC DNA from Streptomyces lividans and Escherichia coli. 638 33

The lack of simple and efficient methods for extraction of DNA from Nocardia spp. has hampered molecular manipulation of the DNA for diagnostic purposes. In the present study, a method for the rapid extraction of undegraded genomic nocardial DNA was established. Briefly, 14 pathogenic Nocardia strains were grown at 37 degrees C for 3 to 5 days in Sauton broth containing 0.05% Tween 80. Subsequently, the cultures were treated for 48 h with 1.2 mg of cycloserine per ml (final concentration). Cells were then harvested by centrifugation and treated with a lysis solution containing 3 mg of lysozyme per ml. This was followed by the addition of proteinase K and sodium dodecyl sulfate to final concentrations of 0.2 mg/ml and 0.5%, respectively, and incubation for 1 h at 50 degrees C. DNA was precipitated with isopropanol after phenol-chloroform-isoamyl alcohol extractions and RNase treated before being quantitated and analyzed by agarose gel electrophoresis. The average undegraded DNA yields obtained were 101 micrograms for Nocardia brasiliensis and 121 micrograms for N. asteroides. This DNA was suitable for restriction endonuclease digestion and PCR amplification, which are methods being applied to the characterization and diagnosis of slowly growing organisms such as Nocardia spp.
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PMID:A rapid and gentle method for isolation of genomic DNA from pathogenic Nocardia spp. 887 44

The isolation of intact, high-molecular-mass genomic DNA is essential for many molecular biology applications including long PCR, endonuclease restriction digestion, Southern blot analysis, and genomic library construction. Many protocols are available for the extraction of DNA from plant material. However, for latex-containing Asteraceae (Cichorioideae) species, standard protocols and commercially available kits do not produce efficient yields of high-quality amplifiable DNA. A cetyltrimethylammonium bromide protocol has been optimized for isolation of genomic DNA from latex-containing plants. Key steps in the modified protocol are the use of etiolated leaf tissue for extraction and an overnight 25 degrees C isopropanol precipitation step. The purified DNA has excellent spectral qualities, is efficiently digested by restriction endonucleases, and is suitable for long-fragment PCR amplification.
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PMID:Extraction of high-quality genomic DNA from latex-containing plants. 1267 15

The potential of a nucleic acid-based optical bioprobe for environmental measurements and drug monitoring is described. The sensor employs the long-wavelength intercalating fluorophore TO-PRO-3 (TP3). Compounds that interact with the TP3-DNA complex are indirectly detected by a decrease in the fluorescence intensity. We found that the configuration and length of the DNA dramatically affected the intensity of the fluorescence emitted from the TP3-DNA complex. We compared nucleic acids from different sources and optimized the system for pBR322 plasmid DNA (4363 bp) digested by HindIII restriction endonuclease. This endonuclease has a single recognition site in plasmid pBR322. In the proposed method, we attempt to combine broad-range detection with rapid and simple operation. A fiber-optic capillary fluorescence system was used to analyze toxic aromatic amines, antibiotics, and several kinds of antitumor drugs, using small amounts of sample, down to 10 muL, with a sensitivity comparable to that of current electrochemical methods. The detection limit can be as low as a few ppb or submicromolar. This approach is useful for routine screening in environmental monitoring or for controlling cytotoxic drug administration. The ease of operation and the rapid response allow high-throughput screening.
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PMID:Fluorometric broad-range screening of compounds with affinity for nucleic acids. 1582 80

Minicircle (MC) DNA vectors are able to generate a high-level transgene expression in vivo, which is superior to the one afforded by conventional plasmids. MC vectors are produced by replicating a parental plasmid (PP) and promoting its recombination in Escherichia coli. This generates a MC with the expression cassette, and a miniplasmid (MP) with the replication segment. Unfortunately, wider use of MC vectors is hampered by difficulties in isolating the target MCs from their MP counterpart. In this proof-of-concept study, a reproducible process is described to improve the purification of supercoiled (sc) MCs that combines an in vitro enzymatic relaxation of sc MP impurities with topoisomer separation and RNA clearance by hydrophobic interaction chromatography (HIC) step. At the early stage of vector design, a site for the nicking endonuclease Nb.BbvCI was strategically placed in the MP part of the PP backbone. A process was then established that involves E. coli culture and recombination of PPs into target MC, cell harvesting and alkaline lysis, precipitation with isopropanol and ammonium sulfate and diafiltration/concentration by microfiltration. Next, an in vitro digestion step was carried out with Nb.BbvCI to nick of one of the strands of the MPs and of non-recombined PPs by Nb.BbvCI. As a result, sc MPs and non-recombined PPs were converted into the corresponding open circular (oc) forms whereas sc MCs remain unaffected. Finally, sc MC was isolated from oc DNA molecules (oc MPs, oc MC) and RNA by performing HIC with a phenyl-Sepharose column using a series of elution steps with decreasing ammonium sulfate concentrations. On the basis of agarose gel electrophoresis analysis, the sc MC-containing fractions were determined to be virtually free from nucleic acid impurities.
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PMID:Development of a nicking endonuclease-assisted method for the purification of minicircles. 2701 16