scholarly journals Physicochemical properties of kinetoplast DNA from Crithidia acanthocephali. Crithidia luciliae, and Trypanosoma lewisi.

1975 ◽  
Vol 67 (2) ◽  
pp. 378-399 ◽  
Author(s):  
D L Fouts ◽  
J E Manning ◽  
D R Wolstenholme
Keyword(s):  
Base Composition ◽  
Unit Length ◽  
Buoyant Density ◽  
Cesium Chloride ◽  
Kinetoplast Dna ◽  
Molar Ratios ◽  
Trypanosoma Lewisi ◽  
Crithidia Luciliae ◽  
Sequence Homologies ◽  
Thermal Melting

The protozoa Crithidia and Trypanosoma contain within a mitochondrion a mass of DNA known as kinetoplast DNA (kDNA) which consists mainly of an association of thousands of small circular molecules of similar size held together by topological interlocking. Using kDNA from Crithidia acanthocephali, Crithidia luciliae, and Trypanosoma lewisi, physicochemical studies have been carried out with intact associations and with fractions of covalently closed single circular molecules, and of open single circular and unit length linear molecules obtained from kDNA associations by sonication, sucrose sedimentation, and cesium chloride-ethidium bromide equilibrium centrifugation. Buoyant density analyses failed to provide evidence for base composition heterogeneity among kDNA molecules within a species. The complementary nucleotide strands of kDNA molecules of all three species had distinct buoyant densities in both alkaline and neutral cesium chloride. For C. acanthocephali kDNA, these buoyant density differences were shown to be a reflection of differences in base composition between the complementary nucleotide strands. The molar ratios of adenine: thymine:guanine:cytosine, obtained from deoxyribonucleotide analyses were 16.8:41.0:28.1:14.1 for the heavy strand and 41.6:16.6:12.8:29.0 for the light strand. Covalently closed single circular molecules of C. acanthocephali (as well as intact kDNA associations of C. acanthocephali and T. lewisi) formed a single band in alkaline cesium chloride gradients, indicating their component nucleotide strands to be alkaline insensitive. Data from buoyant density, base composition, and thermal melting analyses suggested that minor bases are either rare or absent in Crithidia kDNA. The kinetics of renaturation of 32P labeled C. acanthocephali kDNA measured using hydroxyapatite chromatography were consistent with at least 70% of the circular molecules of this DNA having the same nucleotide sequence. Evidence for sequence homologies among the kDNAs of all three species was obtained from buoyant density analyses of DNA in annealed mixtures containing one component kDNA strand from each of two species.

scholarly journals Replication of kinetoplast DNA of Crithidia acanthocephali. I. Density shift experiments using deuterium oxide.

1976 ◽  
Vol 70 (2) ◽  
pp. 406-418 ◽  
Author(s):  
J E Manning ◽  
D R Wolstenholme
Keyword(s):  
Nuclear Dna ◽  
Deuterium Oxide ◽  
Gradient Centrifugation ◽  
Unit Length ◽  
Buoyant Density ◽  
Cesium Chloride ◽  
Kinetoplast Dna ◽  
Open Circular ◽  
Linear Molecules ◽  
Topological Interlocking

The protozoan Crithidia acanthocephali contains, within a modified region of a mitochondrion, a mass of DNA known as kinetoplast DNA (kDNA). This DNA consists mainly of an association of approximately 27,000 covalently closed 0.8-mum circular molecules which are apparently held together in a definite ordered manner by topological interlocking. After culturing of C. acanthocephali cells for 25 generations in medium containing 75% deuterium oxide, both nuclear DNA (rhonative, nondeuterated=1.717 g/cm3) and kDNA (rhonative, nondeuterated=1.702 g/cm3) increased in buoyant density by 0.012 g/cm3. The replication of the two DNAs was studied by cesium chloride buoyant density analysis of DNAs from exponentially growing cells taken at 1.0, 1.4, 2.0, 3.0, and 4.0 cell doublings after transfer of cells from D2O-containing medium into medium containing only normal water. The results obtained from analysis of both native and denatured nuclear DNAs indicate that this DNA replicates semiconservatively. From an analysis of intact associations of kDNA, it appears that this DNA doubles once per generation and that the newly synthesized DNA does not segregate from parental DNA. Fractions of covalently closed single circular molecules and of open circular and unit length linear molecules were obtained from associations of kDNA by sonication, sucrose sedimentation, and cesium chloride-ethidium bromide equilibrium gradient centrifugation. Buoyant density profiles obtained from these fractions indicate that: (a) doubling of the kDNA results from the replication of each circular molecule rather than from repeated replication of a small fraction of the circular molecules; (b) replication of kDNA is semiconservative rather than conservative, but there is recombination between the circles at an undefined time during the cell cycle.

scholarly journals KINETOPLAST DEOXYRIBONUCLEIC ACID OF THE HEMOFLAGELLATE TRYPANOSOMA LEWISI

1970 ◽  
Vol 47 (3) ◽  
pp. 689-702 ◽  
Author(s):  
Hartmut C. Renger ◽  
David R. Wolstenholme
Keyword(s):  
Electron Microscope ◽  
Ethidium Bromide ◽  
Thermal Denaturation ◽  
Deoxyribonucleic Acid ◽  
Cesium Chloride ◽  
Kinetoplast Dna ◽  
Microscope Examination ◽  
Main Band ◽  
Trypanosoma Lewisi ◽  
High Degree

Cesium chloride centrifugation of DNA extracted from cells of blood strain Trypanosoma lewisi revealed a main band, ρ = 1.707, a light satellite, ρ = 1.699, and a heavy satellite, ρ = 1.721. Culture strain T. lewisi DNA comprised only a main band, ρ = 1.711, and a light satellite, ρ = 1.699. DNA isolated from DNase-treated kinetoplast fractions of both the blood and culture strains consisted of only the light satellite DNA. Electron microscope examination of rotary shadowed preparations of lysates revealed that DNA from kinetoplast fractions was mainly in the form of single 0.4 µ circular molecules and large masses of 0.4 µ interlocked circles with which longer, often noncircular molecules were associated. The 0.4 µ circular molecules were mainly in the covalently closed form: they showed a high degree of resistance to thermal denaturation which was lost following sonication; and they banded at a greater density than linear DNA in cesium chloride-ethidium bromide gradients. Interpretation of the large masses of DNA as comprising interlocked covalently closed 0.4 µ circles was supported by the findings that they banded with single circular molecules in cesium chloride-ethidium bromide gradients, and following breakage of some circles by mild sonication, they disappeared and were replaced by molecules made up of low numbers of apparently interlocked 0.4 µ circles. When culture strain cells were grown in the presence of either ethidium bromide or acriflavin, there was a loss of stainable kinetoplast DNA in cytological preparations. There was a parallel loss of light satellite and of circular molecules from DNA extracted from these cells.

DNA BASE COMPOSITION OF CYTOPHAGA MARINOFLAVA N. SP. DETERMINED BY BUOYANT DENSITY MEASUREMENTS IN CESIUM CHLORIDE

1966 ◽  
Vol 12 (6) ◽  
pp. 1099-1103 ◽  
Author(s):  
R. R. Colwell ◽  
R. V. Citarella ◽  
P. K. Chen
Keyword(s):  
Nucleic Acid ◽  
Base Composition ◽  
Marine Bacterium ◽  
Deoxyribonucleic Acid ◽  
Buoyant Density ◽  
Cesium Chloride ◽  
Dna Base Composition ◽  
Density Measurements ◽  
Dna Base ◽  
Relationship Of

A marine bacterium, NCMB 397, host strain for bacteriophages NCMB 384 and 385, has been subjected to taxonoinic analysis. Overall base composition of the highly purified deoxyribonucleic acid was determined and found to be 37 moles % guanine + cytosine. The phenetic and nucleic acid data suggest significant relationship of this strain and members of the genus Cytophaga. A description of Cytophaga marinoflava n. sp. is presented.

The pressure dependence of the buoyant density of DNA as a function of base composition

Biopolymers ◽  
1971 ◽  
Vol 10 (12) ◽  
pp. 2615-2617 ◽  
Author(s):  
William Bauer ◽  
Fred Prindaville ◽  
Jerome Vinograd
Keyword(s):  
Pressure Dependence ◽  
Base Composition ◽  
Buoyant Density

scholarly journals The macromolecular properties of blood-group-specific glycoproteins. Characterization of a series of fractions obtained by density-gradient ultracentrifugation

1974 ◽  
Vol 143 (3) ◽  
pp. 669-679 ◽  
Author(s):  
K. Ramakrishnan Bhaskar ◽  
J. Michael Creeth
Keyword(s):  
Density Gradient ◽  
Blood Group ◽  
Buoyant Density ◽  
Physicochemical Characterization ◽  
Cyst Fluid ◽  
Equilibrium Density ◽  
Density Gradient Ultracentrifugation ◽  
Molecular Weights ◽  
Molar Ratios ◽  
Blood Group Substances

1. Equilibrium density-gradient ultracentrifugation in caesium salts was used in two stages in the isolation and subfractionation of the glycoprotein component from a human ovarian-cyst fluid. The eight main subfractions thus obtained were the subject of detailed physicochemical characterization. 2. The fractions were unimodal in buoyant-density distribution, but had discrete ρ0 values ranging from 1.31 to 1.35. 3. Weight-average molecular weights and sedimentation coefficients decreased regularly with decreasing density of the fraction, whereas the partial specific volumes and selective solvation parameters increased. The latter behaviour correlates well with the increasing peptide content of the lighter fractions. 4. The fractions exhibited a range of analytical composition, although all were within the limits previously observed for blood-group substances of Lea specificity. All fractions had approximately equal Lea activity. The peptide content varied systematically from 7% for the densest fraction to 15% for the lightest, but the relative distributions of the amino acids remained essentially constant throughout the series. In particular, serine plus threonine plus proline made up about 50% of the peptide content of all the fractions. Fucose, galactose and N-acetylglucosamine contents decreased with increasing peptide content of the fractions, but N-acetylgalactosamine and sialic acid exhibited the opposite trend. Molar ratios of N-acetylgalactosamine to the sum of serine and threonine remained essentially constant at 0.8–0.9, implying a high degree of glycosylation of all the molecules, but the ratio of N-acetylglucosamine to N-acetylgalactosamine decreased steadily with increasing peptide content, suggesting the presence of oligosaccharide side chains of various lengths. The results are discussed in terms of the accepted structure of glycoprotein molecules. 5. Experiments on the glycoproteins extracted with phenol from the same cyst fluid have confirmed that equilibrium centrifugation in caesium salts does not remove any non-covalently bound protein nor cause any changes in the tertiary structures of these glycoprotein molecules.

A NON-NUCLEOTIDE POLYMER FOUND IN THE DNA OF THE SAND DOLLAR, ECHINARACHNIUS PARMA: II. PRELIMINARY CHARACTERIZATION

1967 ◽  
Vol 45 (2) ◽  
pp. 281-287 ◽  
Author(s):  
Herbert S. Rosenkranz
Keyword(s):  
Buoyant Density ◽  
Sedimentation Coefficient ◽  
Negative Electrode ◽  
Cesium Chloride ◽  
Amino Sugar ◽  
Electrophoretic Analysis ◽  
Positive Test ◽  
Preliminary Characterization ◽  
Acid Hydrolysate

A preliminary characterization of the non-nucleotidic component present in the DNA of Echinarachnius parma was undertaken. This material has an extremely high sedimentation coefficient (907 S). It contains no deoxyribose and presumably no ribose. After acid hydrolysis it was strongly ninhydrin-positive and also gave positive tests for reducing sugars as well as a slightly positive test for amino sugars. Upon electrophoretic analysis of an acid hydrolysate, three ninhydrinpositive spots were detected. One of these migrated to the negative electrode with a mobility identical with that of galactosamine, the other migrated to the positive electrode, and the third was neutral at pH 6.3. The spot with a mobility identical with that of galactosamine also gave a positive test for amino sugar. The material was not attacked by α-amylase. However, digestion with a crude trypsin preparation resulted in loss of the banding property in gradients of cesium chloride. Exposure to purified trypsin did not completely digest it, but caused an increase in buoyant density.

PCR-based identification of Trypanosoma lewisi and Trypanosoma musculi using maxicircle kinetoplast DNA

Acta Tropica ◽  
2017 ◽  
Vol 171 ◽  
pp. 207-212 ◽  
Author(s):  
Xiao-Kun Hong ◽  
Xuan Zhang ◽  
Octavio Alejandro Fusco ◽  
You-Gen Lan ◽  
Zhao-Rong Lun ◽  
...  
Keyword(s):  
Kinetoplast Dna ◽  
Trypanosoma Lewisi
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