Sedimentation of DNA Released from Chinese Hamster Cells

Under high pH and high salt conditions, Chinese hamster cells lyse and release a DNA-containing material of large molecular weight. With increasing lysis time, a smaller material is resolved from the large one. Relative to T4 DNA, the smaller is estimated to be ~2 × 10⁸ daltons (number average). From a comparison of radiation data, the target size of the larger is about 15 times that of the smaller (probably a lower limit estimate). In addition to concentration of alkali, temperature, and time of lysis, the resolution of the smaller from the larger material is shown to be affected by other factors. Three of these are: fluorescent light exposure during lysis, X-irradiation before lysis, and incorporation of actinomycin D before lysis. All of these treatments result in degradation of the smaller molecules if large enough exposures are used. The sedimentation patterns of both DNA materials are strongly speed dependent. This probably results from changes in molecular conformation and concomitant increases in viscous drag with speed. The speed dependence differs qualitatively for the two materials, an observation which suggests that they differ in ways in addition to size.     

辐射后单个细胞DNA结构变化的定量检测

细胞照射后可产生DNA链断裂、DNA-DNA交联、DNA-蛋白质交联等重要的DNA结构损伤,最终可导致DNA高级结构-DNA超螺旋结构状态的改变,而引发DNA复制、表达等一系列改变.参考国外报导,建立了单细胞电泳法(single cell gel electrophoresis assay),并辅以图象分析技术,可快速检测低达0.1Gy剂量所致DNA结构损伤,并得到了较好的剂量-效应关系,可望成为生物剂量计,用于环境低剂量辐射的监测.     

Proteomic Analysis of Proton Beam Irradiated Human Melanoma Cells

Proton beam irradiation is a form of advanced radiotherapy providing superior distributions of a low LET radiation dose relative to that of photon therapy for the treatment of cancer. Even though this clinical treatment has been developing for several decades, the proton radiobiology critical to the optimization of proton radiotherapy is far from being understood. Proteomic changes were analyzed in human melanoma cells treated with a sublethal dose (3 Gy) of proton beam irradiation. The results were compared with untreated cells. Two-dimensional electrophoresis was performed with mass spectrometry to identify the proteins. At the dose of 3 Gy a minimal slowdown in proliferation rate was seen, as well as some DNA damage. After allowing time for damage repair, the proteomic analysis was performed. In total 17 protein levels were found to significantly (more than 1.5 times) change: 4 downregulated and 13 upregulated. Functionally, they represent four categories: (i) DNA repair and RNA regulation (VCP, MVP, STRAP, FAB-2, Lamine A/C, GAPDH), (ii) cell survival and stress response (STRAP, MCM7, Annexin 7, MVP, Caprin-1, PDCD6, VCP, HSP70), (iii) cell metabolism (TIM, GAPDH, VCP), and (iv) cytoskeleton and motility (Moesin, Actinin 4, FAB-2, Vimentin, Annexin 7, Lamine A/C, Lamine B). A substantial decrease (2.3 x) was seen in the level of vimentin, a marker of epithelial to mesenchymal transition and the metastatic properties of melanoma.     

辐照地被竹突变体DNA多态性分析

利用ISSR分子标记技术对γ-射线(铯137)辐照地被竹突变体植株的基因组DNA变异进行检测和分析,结果表明从100条ISSR引物中筛选得到多态性高、重复性好的引物21条,ISSR的多态性主要表现为扩增片段的增减和扩增片段长度的差异,不同的突变株系中,多态性比率为19.46％ ～ 72.39％,反映了各突变体中遗传物质...     

A Comparison of Toxin Production by Clostridium botulinum Type E in Irradiated and Unirradiated Vacuum Packed Fish

S ummary . The rates of toxin production at 10° by inocula of 10⁵, 10⁴, 10³ and 10² spores of Clostridium botulinum type E in vacuum packed herring, cod and haddock were compared with that of equivalent numbers of spores surviving γ-irradiation at 0.3 Mrad. There was little difference between the rates of toxin production in unirradiated and irradiated fish. More toxin was produced in irradiated herring than in unirradiated, but in haddock the reverse was obtained. In cod about equal amounts of toxin were found without trypsinization; after trypsinization toxin levels were generally higher in irradiated samples.     

Immunogenicity of Irradiated Salmonella typhimurium Cells and Endotoxin

The effects of high doses of radiation (1, 5, or 20 Mrad) on the toxicity, pyrogenicity, and immunogenicity of Salmonella typhimurium cells and endotoxin were studied. Toxicity decreased progressively after exposure to 1, 5, or 20 Mrad. The lethal effect of 1-Mrad exposed cells was greater than that of heat-, acetone-, or alcohol-killed preparations. An amount of 5 Mrad is about a 50% end point in terms of inactivation of the lethal lipopolysaccharide or cell-associated determinants. The fever response to radiation-killed salmonellae decreased between 1- and 20-Mrad exposure. The immunogenicity of 1-Mrad-treated cells usually exceeded that of nonirradiated preparations in mouse-protection tests. With increasing radiation doses, there was a dramatic decrease in, but not an abolition of, immunogenicity. Preparations exposed to 20 Mrad which were nonlethal afforded significant protection. The results are interpreted as a reflection of a dissociation of the primary and secondary toxic determinants of endotoxin after irradiation. The data indicate the potential value of radiation sterilization as a means of production of Salmonella vaccine.     

Sedimentation coefficients of replicating and crosslinked DNA

The sedimentation coefficients of branched and cyclic replicating DNA structures have been calculated taking into account both chain stiffness and excluded volume effects. For branched DNA, the calculated ratio S⁰(α)/S⁰(0) of the sedimentation coefficient at degree of replication α to that at α = 0 increases from 1.000 to 1.331 as α goes from 0 to 1. For cyclic replicating DNA, S⁰(1)/S⁰(0) = 1.517. The considerable difference between these two ratios for α = 1 suggests that the two types of replicating structures may feasibly be distinguished by sedimentation velocity studies, A similar calculation has been made of the sedimentation coefficient of singly cross-linked DNA under denaturing conditions, as a function of placement of the crosslink. In 0.195M Na⁺, the ratio S⁰(α)/S⁰(0) varies from 1.000 at α = 0 to 1.124 at α = 0.5, where α is the fractional distance of the crosslink from the end of the chain.     

Sedimentation Changes of L Cells in a Density Gradient Early After Infection with Vaccinia Virus

By 2 h postinfection, LM cells infected with vaccinia virus show a shift in their distribution when separated on a Ficoll density gradient. This shift is dependent on both time and the multiplicity of infection and is due, at least in part, to an increase in cell size. Those cells which do shift in position in the gradient represent infected members of the population. Physical changes induced in virus-infected cells can be utilized for studying early events in virus replication.     

Sedimentation velocity properties of catenated DNA molecules from HeLa cell mitochondria

Catenated molecules of closed circular DNA have been isolated from the mitochondrial DNA of HeLs cells. The sedimentation coefficients of several purified species have been investigated. The catenated dimer, made up of two interlocked duplex circles, sediments at 51 S in its superhelical (closed) form. Treatment with pancreatic DNase to relax the duplex circles converts the 51 S doubly closed dimer to a 42 S singly open species, then to a 36 S doubly open catenated dimer. The triply closed trimer sediments at 63 S and is converted to a 45 S triply open form by DNase. Electron microscopy of the DNA samples before and after DNase treatment shows that under the conditions used DNase does not change the catenated nature of the DNA. The measured sedimentation coefficients, have been compared with those estimated from previously proposed correlations of sedimentation coefficient and molecular weight, and with the sedimentation coefficients for catenated DNA presented by Wang. When all the interlocked circles in a catenane are relaxed, the DNA sediments about 5–10% faster than a relaxed multiple-length circular molecule of the same molecular weight. The sedimentation coefficient, 36 S, of the fully relaxed catenated dimer is 1.4 times that of the relaxed monomer.     

Sedimentation coefficient of polyoma virus DNA

The sedimentation coefficient of the twisted circular form of polyoma virus DNA is calculated from the Kirkwood sedimentation–diffusion equation, the structure being assumed to be a rigid double superhelix. Agreement with the experimental sedimentation coefficient can be obtained, with the use of an experimental value for the number of superhelical turns, when the pitch of the superhelix is intermediate between its minimal and maximal possible values. Another model, which has been proposed for polyoma DNA at low ionic strengths, may be visualized as a superhelical structure wound about a torus. Calculations of sedimentation coefficients for this model agree qualitatively with experimental data at ionic strengths Below 10^?2M.     

Sedimentation coefficient of T-even bacteriophage DNA

The sedimentation coefficient of T2 phage DNA has been studied by zone centrifugation in sector-shaped preparative centrifuge tubes over a concentration range of 0.02–2.0 μg./ml. DNA. These results have been compared to a similar study in the analytical centrifuge of T4 DNA over the range of 0.50–5.75 μg./ml. DNA. A value for the sedimentation coefficient of 60.7 ± 1.8 S. was obtained by the first method and a value of 61.3 ± 1.5 S. by the second.     

DNA Polymerases from Human Cells

Nuclei of HeLa cells contain two separable DNA polymerases which have different properties and cytoplasm contains a single DNA polymerase activity similar to one of the nuclear DNA polymerases. The same distribution of DNA polymerases has been found in the nucleus and cytoplasm of WI-38 cells, a normal diploid human cell line.     

Enhanced UV Sensitivity of Thiobacillus ferrooxidans Resulting from Caffeine and Acriflavine Treatment of Irradiated Cells

This study was aimed at identifying the roles of caffeine and acriflavine, two repair inhibitors, on UV sensitivity of iron-oxidizing Thiobacillus ferrooxidans ATCC 13728. The UV-dose response survival curve was inflected in nature, suggesting the population heterogeneity of the isolate. Caffeine and acriflavine potentiated the UV-induced killing of the organism. With the increase in concentrations of these compounds, the extent of survival decreased. Similarly, the inhibitory effects of caffeine and acriflavine increased with the increase in dose of UV-irradiation. The cells irradiated with 10 s (equivalent to 5.6 × 10⁻⁵ J/m²/s) of UV-exposure tended to become resistant to the inhibitory effects of caffeine and acriflavine, as evidenced by the time course study of recovery. The cells appear to stage a dramatic recovery from UV damage in the presence of caffeine (3.0 mg/ml) and acriflavine (20 μg/ml) over a period of 25–30h and 35–40h respectively, when grown in the presence of energy sources. Received: 4 December 2000/Accepted: 10 January 2001     

Sedimentation of homogeneous double-strand DNA molecules.

Sedimentation velocity studies have been carried out with isolated double-strand DNA fragments prepared by digestion of PM2 phage with the restriction endonuclease Hae III. The results show that DNA molecules shorter than about 200 base pairs behave almost exactly as rigid rods with a diameter of 27 A. The behavior of the larger fragments (up to 1735 base pairs) can be described very well by either the theory of Yamakawa and Fujii (Yamakawa, H., and Fujii, M. (1973), Macromolecules 6. 407) using the same diameter and a persistence length of 575 A, or the theory of Hearst and Stockmayer (Hearst, J.E., and Stockmayer, W.H. (1962), J. Chem. Phys. 37, 1425) using a diameter of 20 A and a persistence length of 525 A.