Continuous DNA replication in the nucleus of the dinoflagellate Prorocentrum micans (Ehrenberg)

1977 ◽  
Vol 27 (1) ◽  
pp. 81-90
Author(s):  
S.A. Filfilan ◽  
D.C. Sigee
Keyword(s):  
Electron Microscope ◽  
Dna Synthesis ◽  
Nuclear Dna ◽  
Continuous Process ◽  
Prorocentrum Micans ◽  
Interphase Nuclei ◽  
Electron Microscope Autoradiography ◽  
Dividing Cells ◽  
High Degree ◽  
Very High

The uptake of tritiated thymine into cells of a heterogeneous population of Prorocentrum micans was investigated using light-microscope and electron-microscope autoradiography. Specificity of thymine uptake into DNA was demonstrated by the specific removal of label from wax-embedded material using DNase and by the high degree of localization of nuclear label to chromosomes in the electron-microscope autoradiographs. All nuclei, including both dividing and non-dividing cells, showed a substantial uptake of label, indicating that nuclear DNA synthesis in Prorocentrum micans is a continuous process. The level of DNA synthesis does show considerable variation, however, with very high levels in some interphase nuclei. The continuous replication of nuclear DNA provides further evidence of dinoflagellate affinity to the prokaryotes, and indicates that Prorocentrum micans is a very primitive eukaryote cell.

scholarly journals CYTOPLASMIC DNA SYNTHESIS IN AMOEBA PROTEUS

1962 ◽  
Vol 15 (3) ◽  
pp. 525-534 ◽  
Author(s):  
M. Rabinovitch ◽  
W. Plaut
Keyword(s):  
Dna Synthesis ◽  
Tritiated Thymidine ◽  
Amoeba Proteus ◽  
Living Cells ◽  
Large Numbers ◽  
Cytoplasmic Dna ◽  
Basic Dyes ◽  
Nuclease Digestion ◽  
High Degree ◽  
Very High

The incorporation of tritiated thymidine in Amoeba proteus was reinvestigated in order to see if it could be associated with microscopically detectable structures. Staining experiments with basic dyes, including the fluorochrome acridine orange, revealed the presence of large numbers of 0.3 to 0.5 µ particles in the cytoplasm of all cells studied. The effect of nuclease digestion on the dye affinity of the particles suggests that they contain DNA as well as RNA. Centrifugation of living cells at 10,000 g leads to the sedimentation of the particles in the centrifugal third of the ameba near the nucleus. Analysis of centrifuged cells which had been incubated with H3-thymidine showed a very high degree of correlation between the location of the nucleic acid-containing granules and that of acid-insoluble, deoxyribonuclease-sensitive labeled molecules and leads to the conclusion that cytoplasmic DNA synthesis in Amoeba proteus occurs in association with these particles.

Timing of nucleolar DNA replication in Amoeba proteus

1976 ◽  
Vol 22 (3) ◽  
pp. 521-530
Author(s):  
I. Minassian ◽  
L.G. Bell
Keyword(s):  
Electron Microscope ◽  
Dna Replication ◽  
Dna Synthesis ◽  
Central Region ◽  
Thymidine Incorporation ◽  
Tritiated Thymidine ◽  
Amoeba Proteus ◽  
Electron Microscope Autoradiography ◽  
Microscope Autoradiography ◽  
G2 Phase

Light- and electron-microscope autoradiography have been used to follow the incorporation of [3H]thymidine at different stages during the interphase of synchronously growing populations of Amoeba proteus. Two main patterns were found for tritiated thymidine incorporation, i.e. DNA synthesis. The major incorporation was in the central region of the nucleus, but a lesser degree of incorporation occurred in the nucleolar region. The bulk of this nucleolar DNA was found to be late replicating, i.e. it replicated during the G2 phase.

scholarly journals CYTOPLASMIC DNA SYNTHESIS IN AMOEBA PROTEUS

1964 ◽  
Vol 22 (3) ◽  
pp. 505-513 ◽  
Author(s):  
D. R. Wolstenholme ◽  
W. Plaut
Keyword(s):  
Electron Microscope ◽  
Dna Synthesis ◽  
Light Microscope ◽  
Tritiated Thymidine ◽  
Amoeba Proteus ◽  
Electron Microscope Autoradiography ◽  
Cytoplasmic Dna ◽  
Microscope Autoradiography ◽  
Normal Particle ◽  
Labeling Pattern

The application of electron microscope autoradiography to Amoeba proteus cells labeled with tritiated thymidine has permitted the identification of morphologically distinct particles in the cytoplasm as the sites of incorporated DNA precursor. The particles correspond to those previously described from light microscope studies, with respect to both H3Tdr incorporation and distribution in centrifugally stratified amoebae. Ingested bacteria differ from the particles, in morphology as well as in the absence of associated label. Attempts to introduce a normal particle labeling pattern by incubating amoebae with labeled sediment derived from used amoeba medium failed. The resultant conclusion, that the particles are maintained in the amoeba by self-duplication, is supported by the presence of particles in configurations suggestive of division.

scholarly journals Microbubbles in replicating nuclear deoxyribonucleic acid from Physarum polycephalum

1979 ◽  
Vol 183 (2) ◽  
pp. 477-480 ◽  
Author(s):  
D A F Gillespie ◽  
N Hardman
Keyword(s):  
Electron Microscopy ◽  
Electron Microscope ◽  
Dna Synthesis ◽  
Physarum Polycephalum ◽  
Deoxyribonucleic Acid ◽  
Nuclear Dna ◽  
S Phase ◽  
Dna Structures ◽  
Nuclear Deoxyribonucleic Acid

Clusters of microbubbles, represent probable sites of newly initiated DNA synthesis, were identified in nuclear DNA from Physarum polycephalum by using the electron microscope. Their presence is associated specifically with S-phase. Each microbubble corresponds in size to a replicating segment of DNA about 100-5000 nucleotide residues in length. The DNA structures containing microbubbles are metastable, and revert to native DNA in the presence of moderate concentrations of formamide used to prepare samples for electron microscopy. It is suggested that each cluster of microbubbles may correspond to a unit of replication (a replicon) in Physarum DNA.

scholarly journals High-Speed Electron Microscope Autoradiographic Studies of Diffusible Compounds,

1981 ◽  
Vol 29 (1A_suppl) ◽  
pp. 143-160 ◽  
Author(s):  
Vinci Mizuhira ◽  
Michiko Shiihashi ◽  
Yutaka Futaesaku
Keyword(s):  
Electron Microscope ◽  
Exposure Time ◽  
High Speed ◽  
Nuclear Dna ◽  
Specific Activity ◽  
High Specific Activity ◽  
Good Resolution ◽  
Cell Organelles ◽  
Electron Microscope Autoradiography ◽  
Resolution Problem

Three important factors are necessary for successful electron microscope autoradiography (EM-ARG): good resolution, proper preparation of the radioactive isotope (RI) labeled diffusible compounds, and shortened exposure time for ARG. The resolution problem is fundamental to EM-ARG. However, unless the diffusible RI compounds have been fixed correctly in the tissues during preparation, good resolution is useless. It is also necessary to shorten the exposure time for ARG. As yet, a high-speed ARG method for electron microscopy has not been reported, although scintillation ARG methods have been applied to macro- and micro-ARG since 1960. High specific activity, a large amount of radioactivity per unit exposure for radio incorporation (incubation), and careful selection of labeled compounds that concentrate in the DNA or RNA of cell organelles may increase the sensitivity of the emulsion and shorten the exposure time for ARG. For example, labeled thymidine accumulates in nuclear DNA, 3H-SPG (Schizophyllan-produced polyglucan) is incorporated into lysosomal granules, and labeled iodine concentrates in thyroid follicles, often increasing the sensitivity of the emulsion and shortening the exposure time. High-speed ARG yields good data in a very short time, but high-resolution ARG continues to be necessary, even though it requires 4 weeks or more of exposure time. Scintillation autoradiography using tritium seems unstable. We propose a new way to shorten exposure time for EM-ARG, by combining overdevelopment with coating both sides of the grid with emulsion. This method is approximately 100 times more sensitive than the conventional method, and only 4 days of exposure time are required, in contrast to the 1 month usually needed.

scholarly journals Molecular profile of the D. melanogaster mutant genotype w1118 in the presence of variable amount of deuterium (D)

2021 ◽  
Author(s):  
Gallia Butnaru ◽  
◽  
Sorina Popescu ◽  
Keyword(s):  
Drosophila Melanogaster ◽  
Dna Synthesis ◽  
Nuclear Dna ◽  
Culture Media ◽  
Mutant Line ◽  
Molecular Profile ◽  
Variable Amount ◽  
Mutant Genotype ◽  
Very High

The Drosophila melanogaster w1118 mutant line was used to identify the effect of deuterium (D) on DNA synthesis. D concentrations ranged from 30ppm to 96.89% (low and very high amount respec-tively). Five generations of flies were bred on culture media prepared with 6 concentrations of D. For each generation the DNA was analyzed, and its variability was established. The results showed a small involvement of D in the successive synthesis of nuclear DNA.

Continuous Nucleolar DNA Synthesis in Late-Interphase Nuclei of Physarum Polycephalum after Transplantation into Postmitotic Plasmodia

1974 ◽  
Vol 15 (1) ◽  
pp. 131-143
Author(s):  
E. GUTTES
Keyword(s):  
Dna Replication ◽  
Dna Synthesis ◽  
Physarum Polycephalum ◽  
Nuclear Dna ◽  
S Phase ◽  
Interphase Nuclei ◽  
G2 Phase

In the myxomycete, Physarum polycephalum, nuclear DNA synthesis commences immediately upon completion of mitosis. While the synthesis of extranucleolar DNA is completed within a few hours, nucleolar DNA synthesis occurs during most of the S-phase and the entire G2 phase of the intermitotic period. When large (polyploid), late-interphase nuclei were allowed to bypass mitosis by transplantation into recipient plasmodia which were at early interphase and which belonged to a strain having smaller nuclei, the nucleolar DNA of the transplanted nuclei continued to be labelled (autoradiographs) after incubation of the host plasmodia with [3H]thymidine until they entered prophase along with the nuclei of the host plasmodium, approximately one intermitotic period later. This labelling was DNase-sensitive and RNase-resistant. When late-interphase nuclei were labelled with [3H]thymidine just prior to transplantation, there was no decrease of label after transplantation during the additional intermitotic period. We conclude from these experiments that there is no obligatory alternation between nucleolar DNA duplication and mitosis in Physarum polycephalum and that nucleolar DNA replication might exhibit amplification during an experimentally prolonged intermitotic period.

scholarly journals CHROMOSOMAL DNA SYNTHESIS IN DROSOPHILA MELANOGASTER

1968 ◽  
Vol 39 (2) ◽  
pp. 415-429 ◽  
Author(s):  
E. F. Howard ◽  
W. Plaut
Keyword(s):  
Drosophila Melanogaster ◽  
Dna Synthesis ◽  
Chromosome Region ◽  
Chromosomal Dna ◽  
Ordered Arrays ◽  
Intermediate Time ◽  
High Degree ◽  
Chromosome Segments ◽  
Very High ◽  
Chromosome Regions

Analysis of labeling patterns in three chromosome segments of Drosophila melanogaster has shown that the replicative activity within chromosomes is temporally ordered. Moreover, specific labeling patterns on one chromosome occur with specific patterns on another chromosome with a very high degree of correlation. This circumstance leads to the conclusion that DNA synthesis among all the regions in the three chromosome segments studied is coordinated. The various labeling patterns observed in any one chromosome and the combinations of labeling patterns observed in all three chromosome segments can be arranged in ordered arrays, if one assumes that the DNA synthesis in each chromosome region will go to completion without stopping once it has started. Such arrays can serve as models for the temporal order of DNA synthesis among chromosome regions. They predict that in any one chromosome DNA replication begins and ends at very few loci and that synthesis at a larger number of points occurs at an intermediate time.

Electron Microscopy in Molecular Biology

1967 ◽  
Vol 25 ◽  
pp. 2-3
Author(s):  
Cecil E. Hall
Keyword(s):  
Electron Microscopy ◽  
Molecular Biology ◽  
Noise Level ◽  
Molecular Structures ◽  
Material Structure ◽  
The Arts ◽  
Shadow Casting ◽  
Electron Image ◽  
High Degree ◽  
Very High

The visualization of organic macromolecules such as proteins, nucleic acids, viruses and virus components has reached its high degree of effectiveness owing to refinements and reliability of instruments and to the invention of methods for enhancing the structure of these materials within the electron image. The latter techniques have been most important because what can be seen depends upon the molecular and atomic character of the object as modified which is rarely evident in the pristine material. Structure may thus be displayed by the arts of positive and negative staining, shadow casting, replication and other techniques. Enhancement of contrast, which delineates bounds of isolated macromolecules has been effected progressively over the years as illustrated in Figs. 1, 2, 3 and 4 by these methods. We now look to the future wondering what other visions are waiting to be seen. The instrument designers will need to exact from the arts of fabrication the performance that theory has prescribed as well as methods for phase and interference contrast with explorations of the potentialities of very high and very low voltages. Chemistry must play an increasingly important part in future progress by providing specific stain molecules of high visibility, substrates of vanishing “noise” level and means for preservation of molecular structures that usually exist in a solvated condition.

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