Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: 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 physical location of histone molecules in a simian virus 40 DNA-histone complex isolated from purified virions was examined using site-specific restriction endonucleases. The complex contains four host histone species but lacks histone F1. Histones prevent complete cleavage of SV40 DNA by two restriction enzymes, HindIII and EcoRI. From the pattern of DNA fragments resulting from cleavage of the histone-DNA complex by the HindIII endonuclease, which makes six breaks on purified SV 40 DNA, we have concluded that histones are randomly arranged on SV40 DNA relative to restriction enzyme cleavage sites. The EcoRI endonuclease, which makes one break in SV40 NDA, was used to determine the degree of physical coverage of the SV 40 DNA molecule by histones. We observed that 80% of the EcoRI sites in the complex are accessible to the enzyme while 20% are "closed." This degree of coverage is consistent with the mass ratio of DNA:histone in the complex as revealed by the buoyant density of the formaldehyde-fixed complex. We conclude that the histones in the complex are located randomly on the SV 40 genome and cover approximatley 20% of the DNA. These results suggest that the histone species F2b, F2al, F2a2, and F3 are bound without regard to nucleotide sequence of SV 40 DNA.
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PMID:Location of histones on simian virus 40 DNA. 17 47

Isolated SV40 minichromosomes [1-3] were treated with different single-cut restriction endonucleases to probe the arrangement of nucleosomes in relation to the SV70 DNA sequence. While Eco RI and Bam HI each cut 22-27% of the SV40 minichromosomes under limit-digest conditions, Bgl I, which cuts SV40 DNA at or very near the origin of replication [4,5], cleaves 90-95% of the minichromosomes in a preparation. Similar results were obtained with minichromosomes which had been fixed with formaldehyde before endonuclease treatment. One possible interpretation of these findings is that the arrangement of nucleosomes in the compact SV40 minichromosomes is nonrandom at least with regard to sequences near the origin of DNA replication.
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PMID:SV40 viral minichromosome: preferential exposure of the origin of replication as probed by restriction endonucleases. 21 58

Superhelical covalently closed circular replicative form DNA (RF I) of coliphage M13 appears as a relaxed molecule that has a base-unpaired region in the form of a bubble (100 to 200 base pairs long) seen in electron micrographs when spread in the presence of formaldehyde and formamide or after pretreatment with glyoxal. S1 endonuclease, specific for single-stranded DNA, converts superhelical M13 RF I DNA, but not nonsuperhelical M13 RF I to a significant extent, into unit-length linear molecules by sequential nicking of two strands. The locations of S1 nuclease-susceptible sites and glyoxal-fixed base-unpaired regions were both related to the five A-T-rich regions in M13 RF DNA. While S1 nuclease does not show preference for any of these sites, glyoxal-fixed bubbles occur predominantly at the major A-T-rich region in M13 RF DNA.
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PMID:Base-unpaired regions in supercoiled replicative form DNA of coliphage M13. 32 5

Rat liver DNA may be separated into two fractions by stepwise elution from benzoylated-DEAE-cellulose with NaCl and caffeine solutions respectively. Other studies using bacterical and yeast DNA suggested that the first fraction contains native DNA, whereas the second may exhibit some degree of single-stranded character. In the present experiments, chromatography of DNA was monitored by labelling in vivo with [methyl-3H]thymidine in rats previously subjected to partial hepatectomy. In animals killed up to 1 h after thymidine injection, radioactivity eluted in the second fraction was inversely related to the incorporation time, being greatest when animals were killed 10 min after radioisotope injection. However, for most experiments, animals were allowed to survive 2-4 weeks after surgery before use, analysis being made on non-dividing DNA. Under these conditions, the proportion of caffeine-eluted DNA was decreased by subjecting the preparation to shear, before chromatography. A procedure that resulted in 12% of the recovered radioactivity being eluted with caffeine was adopted for experiments involving comparisons of the two DNA fractions. Under these conditions, cross-contamination could be detected by rechromatography, but this did not preclude distinction being made between the two fractions in terms of DNA structure. NaCl-eluted DNA did not bind to nitrocellulose filters. Caffeine-eluted DNA was retained by the filters and released by washing with 3mM-Tris/HCl,pH9.4. The fractions did not differ in terms of isopycnic centrifugation in CsCl. The NaCl-eluted fraction migrated as a single band in polyacrylamide gels, and this pattern was not modified by prior digestion with Neurospora crassa endonuclease. In contrast, caffeine-eluted DNA contained a minor component having a wide molecular-weight distribution and was subject to limited digestion by the endonuclease. The kinetics of denaturation of NaCi-eluted DNA in the presence of formaldehyde, in common with unfractionated DNA, were consistent with double-stranded structure. The same analysis of caffeine-eluted DNA revealed structural abnormality equivalent to two defects per 10000 base-pairs. The data are consistent with the minor fraction of rat liver DNA, separated by using benzoylated-DEAE-cellulose, containing regions of local denaturation. We previously showed that administration of the hepatocarcinogen dimethylnitrosamine is associated with an increase in the proportion of caffeine-eluted DNA. In terms of most analysis, differences between DNA fraction from nitrosamine-treated rats were similar to differences exhibited by preparations from control animals. However, structural analysis using denaturation kinetics indicated defects in both the NaCl- and caffeine-eluted DNA isolated from nitrosamine-treated rats. The two fractions differed from each other in that caffeine-eluted DNA exhibited a degree of structural damage far greater than that detected in any preparation from control animals...
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PMID:Structural defects in rat liver deoxyribonucleic acid. Endogenous single-strained regions in comparison with damage induced in vivo by a carcinogen. 48 85

A formaldehyde denaturation map of the replicative form of phiX174 DNA is obtained. The RFI DNA was converted into a linear state by restriction endonuclease pst I which introduces into this DNA a single double-stranded break. The map has four clear-cut peaks. Their positions excellently correlate with the peak positions on the map of equilibrium denaturation theoretically obtained earlier from the known nucleotide sequence of phiX174 DNA. The sequence is also used for a calculation of the maps of smoothed AT-content. The maxima on these maps correlate well with the peaks on the denaturation maps. To reveal the causes of a good correlation between the experimental formaldehyde and theoretical equilibrium denaturation maps, the theoretical formaldehyde denaturation maps are calculated for different conditions (temperature, formaldehyde concentration) using the detailed theory of DNA interaction with formaldehyde developed earlier.
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PMID:Denaturation maps of DNA: experimental and theoretical maps of phiX174 DNA. 56 10

Mitochondrial DNA (mt DNA) from a patulin producer, Penicillium urticae (synonym P. griseofulvum), was 27.8 kb +/- 0.6 kb in size by electron microscopy and 27.2 kb by agarose gel electrophoresis. Restriction endonuclease maps for nine restriction enzymes were constructed, and eleven fragments which covered the total range of the mt DNA were cloned into the Escherichia coli plasmid vector pUC19. Southern analysis of the native genomes of P. urticae and P. chrysogenum with six of the cloned fragments as probes indicated similar genome arrangements as well as similar restriction maps. Both the large and small rRNA genes of P. urticae and P. chrysogenum were located on these restriction maps using Southern hybridization, and the result also supported the similar arrangement. Agarose/formaldehyde gel electrophoresis indicated that the small rRNA was 1.5 kb in size in both species; but, surprisingly, the large rRNA was 4.2 kb in size for P. urticae and 3.5 kb for P. chrysogenum. These sizes were, respectively, 1.1 kb and 0.4 kb larger than those from the very closely related Aspergillus nidulans.
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PMID:Characterization and comparison of mitochondrial DNAs and rRNAs from Penicillium urticae and P. chrysogenum. 169 25

The photochemically derived silver stain of nucleic acids in polyacrylamide gels originally described by Merril et al. (1981, Science 211, 1437-1438) was modified to reduce unspecific background staining and increase sensitivity (down to 1 pg/mm2 band cross-section). Detection limits for double-stranded DNA fragments from HaeIII endonuclease digests of phage phi X174 were maintained despite eliminating oxidation pretreatment of fixed gels and reducing silver nitrate concentration. Preexposure to formaldehyde during silver impregnation enhanced sensitivity and the inclusion of the silver-complexing agent sodium thiosulphate in the image developer decreased background staining. Higher formaldehyde concentration during image development resulted in darker bands with good contrast. The procedure almost halves the number of steps, solutions and experimental time required and can be used for the staining of DNA fragments in polyacrylamide gels bound to a polyester backing film by controlling temperature during image development. We have applied this improved staining procedure for the routine analysis of complex DNA profiles generated by DNA amplification fingerprinting (DAF).
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PMID:Fast and sensitive silver staining of DNA in polyacrylamide gels. 171 76

The mechanism of DNA degradation and its clinical applications were examined. When purified lambda phage and extracted liver DNA were fixed in phosphate buffered formaldehyde, the DNA did not degrade, but there was incomplete digestion with endonuclease. Rat liver tissues were fixed under various conditions and DNA extracted. Immediate fixation with buffered formaldehyde at low temperature, or the addition of EDTA to buffered formaldehyde blocked the DNA degradation. Analysis of pulsed field gel electrophoresis also showed that DNA was degraded before extraction. These results suggest that tissue nuclease has an important role in DNA degradation in tissue. Furthermore, formaldehyde fixation at low temperature, which may take time and which decreases slightly the staining capacity, is useful for the extraction of intact DNA. For clinical application, the detection of provirus was examined. Genomic DNA was extracted from a necropsy sample of adult T cell leukaemia fixed in formaldehyde; human T cell leukaemia virus type-I (HTLV-I) provirus was successfully detected by Southern blotting.
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PMID:Fundamental study on the mechanism of DNA degradation in tissues fixed in formaldehyde. 212 Feb 90

A dot blot hybridization assay was developed for use as a rapid screening test to detect bovine viral diarrhea virus (BVDV) in serum from infected cattle. A 1.1. kilobase cDNA, prepared from the BVDV genome, was molecularly cloned and used in this study. Insert cDNA was removed from the pUC9 plasmid vector by Pst-I restriction endonuclease digestion and purified from plasmid DNA by agarose electrophoresis and electroelution. The hybridization probe was prepared by nick translation in the presence of gamma dCT32P and labelled to a specific activity of 2 x 10(8) cpm/micrograms of DNA. Specificity was determined by dot blot hybridization of infected cell culture supernate from nine different BVDV strains. The probe hybridized equally with all strains of BVDV tested, which included four cytopathic and five noncytopathic strains of BVDV. Serum was collected from veal calves with respiratory tract disease, unthriftiness, anorexia, and/or poor conditions. Serum samples were treated with nonidet P40 detergent and denatured with formaldehyde and heat prior to application on 1.2 micron nylon membrane filters using a vacuum dot blot apparatus. Hybridization was done under relatively stringent conditions (50% formamide at 42 degrees C). A total of 141 serum samples from different calves were tested and of these samples, 55 (39%) were positive by dot blot hybridization for BVDV RNA. Eight calves (33%) out of 24, tested 3 to 4 weeks later, remained positive for BVDV RNA.
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PMID:Detection of bovine viral diarrhea virus in serum from cattle by dot blot hybridization assay. 217 27

Inactivation of foot-and-mouth disease virus (FMDV) by means of virion-associated endonuclease was found to be suited to the production of safe and potent vaccines, which proved to be equal or better than those containing formaldehyde or ethyleneimine in guinea-pig potency tests. First order inactivation kinetics were regularly shown, with half life values which varied according to the different temperatures used. Inactivation brought about extensive degradation of FMDV RNA, while it did not adversely influence the integrity of critical viral epitopes on FMDV VP1.
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PMID:Safety and efficacy of foot-and-mouth disease vaccines containing endonuclease-inactivated virions. 282 95


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