Detection of pathogenic bacteria by using zinc finger protein fused with firefly luciferase

We constructed a novel bacterial genome detection system using zinc finger protein (ZF) fused with firefly luciferase (ZF-luciferase). Taking advantage of the direct recognition of double-stranded DNA (dsDNA) by ZF, we previously constructed bacterial genome detection systems that did not require dehybridization processes. To detect polymerase chain reaction (PCR) products rapidly and with a high sensitivity, we constructed two kinds of ZF-luciferase, Sp1-fused luciferase (Sp1-luciferase), and Zif268-fused luciferase (Zif268-luciferase). ZF-luciferase not only maintains luciferase activity but also shows dsDNA-binding ability and specificity. Furthermore, we succeeded in the detection of 10 copies of the genome of Legionella pneumophila and Escherichia coli O157. ZF-luciferase would be a useful tool for highly sensitive detection of pathogenic bacterial genome.     

Non-inductive start up of QUEST plasma by RF power

Both start-up and sustainment of plasma were successfully achieved by fully non-inductive current drive using microwave with a frequency of 8.2 GHz. Plasmas current of 15 kA was implemented for 1 s. Magnetic surface reconstruction exhibited a plasma shape with an aspect ratio of below 1.5. The plasma current was dependent significantly on the launchen microwave power and vertical magnetic field, whiile not affected by the mode of launched wave and the toroidal refractive index. Hard X-ray (HXR) emitted from energetic electrons accelerated by the microwave was observed, and the discharge with a plasma current over 4 kA followed the same trend as the number of photons of 10 to 12 keV. This suggests that the plasma current may be driven by energetic electrons. Based on the experimental conditions, alternative explanations of how the plasma current could be driven are discussed.     

Inhibitory Effects of Cholesterol Derivatives on DNA Polymerase and Topoisomerase Activities, and Human Cancer Cell Growth

This paper describes the inhibitory activities of cholesterol derivatives such as cholesterol, sodium cholesteryl sulfate, cholesteryl-5α, 6α-epoxide, cholesteryl chloride, cholesteryl bromide, and cholesteryl hemisuccinate (compounds 1–6, respectively) against DNA polymerase (pol), DNA topoisomerase (topo), and human cancer cell growth. Among the compounds tested, compounds 2 and 6 revealed themselves to be potent inhibitors of animal pols, and the IC₅₀ values for pols were 0.84–11.6 and 2.9–148 μM, respectively. Compounds 2, 3 and 6 inhibited the activity of human topo II, with IC₅₀ values of 5.0, 12.5 and 120 μM, respectively. Compounds 2, 3 and 6 also suppressed human cancer cell (promyelocytic leukemia cell line, HL-60) growth, and LD₅₀ values were 8.8, 20.2 and 72.3 μM, respectively, suggesting that cell growth inhibition had the same tendency as the inhibition of topos rather than pols. Compounds 2 and 6 arrested the cells in S and G2/M phases, compound 3 arrested the cells in the G2/M phase, and these compounds also increased sub-G1 phase in the cell cycle. These results suggested that the effect of cell cycle arrest might be effective on both pols and topos activities. From these findings, the action mode of cholesterol derivatives as anti-cancer compounds is discussed.     

北京养老服务机构入住理由及位置选择的初探——关于合理布局建设养老服务机构

通过对北京市养老服务机构的入住老人进行问卷调研,解析老人的入住理由。老人在机构选择意向中重视"离子女居住地近"的要素,并在实际选择入住时,呈现选择离子女居住地较近机构的趋势。根据该倾向,研究通过整理各功能区特征,为解决北京市养老服务机构入住率分布不均衡的现状,提出各功能区养老机构的规划建设见解。     

Quantitative bioimaging analysis of gonads in olvas-GFP/ST-II YI Medaka (transgenic Oryzias latipes) exposed to ethinylestradiol

This study investigates the adverse and persistent effects of ethinylestradiol (EE2) on mature gonads of transgenic olvas-GFPIST II-YI medaka (Oryzias latipes). The measurement of gonadal size calculating the GFP-fluorescent area was used as a technique that enabled monitoring gonads in living specimens by GFP fluorescence. First, mature medaka were exposed to EE2 (47.8-522 ng/L) for 4 weeks. The gonads showed a significant reduction of the GFP-fluorescent area and Gonadosomatic Index in males exposed to EE2 at >216 ng/L and females exposed at 522 ng/L. Histologically, males at all treatments exhibited testis-ova and additionally, high connective tissue prevalence at > or =216 ng/L. Next, mature male medaka were exposed to EE2 (43.7-473 ng/L) for 3 weeks and allowed to depurate for 6 weeks, to investigate persistent effects of EE2. Continuous gonad observation showed that GFP began to decline 3 weeks after initial exposure to > or =215 ng/L. After depuration, the gonad's fluorescent areas gradually recovered, with no statistical difference at the end of the depuration period; normal spermatogenesis was present in these individuals. Alterations in GFP fluorescence clearly indicate the condition of the gonad in transgenic medaka and this strain showed a facilitated screening fish model to detect the adverse effects on the gonad by estrogenic chemicals.     

Development of a genetic transformation system using new selectable markers for fission yeast Schizosaccharomyces pombe

We describe the development of a new transformation system, using multiple auxotrophic marker genes, for the fission yeast Schizosaccharomyces pombe. We developed three new auxotrophic marker genes (arg12(+), tyr1(+) and ade7(+)) and generated a new host strain, YF043, by Cre-loxP-mediated gene disruption. YF043 possessed six mutated biosynthetic genes (leu1-32, ura4-M190T, arg12::loxP, tyr1::loxP, ade7::loxP and his2::loxP). The combination of this host strain and the new selectable markers can be used for gene disruption using the same preexisting transformation systems. In addition, Sz. pombe vectors were constructed, containing selectable marker genes that complement the auxotrophies of YF043. These new vectors are available for gene disruption and heterologous protein expression in strain YF043. The new Sz. pombe host strain will be a useful tool for molecular genetic studies of Sz. pombe where multiple recombinant modifications or multiple mutations are needed.     

Homology-Based Interactions between Small RNAs and Their Targets Control Dominance Hierarchy of Male Determinant Alleles of Self-Incompatibility in Arabidopsis lyrata

Self-incompatibility (SI) is conserved among members of the Brassicaceae plant family. This trait is controlled epigenetically by the dominance hierarchy of the male determinant alleles. We previously demonstrated that a single small RNA (sRNA) gene is sufficient to control the linear dominance hierarchy in Brassica rapa and proposed a model in which a homology-based interaction between sRNAs and target sites controls the complicated dominance hierarchy of male SI determinants. In Arabidopsis halleri, male dominance hierarchy is reported to have arisen from multiple networks of sRNA target gains and losses. Despite these findings, it remains unknown whether the molecular mechanism underlying the dominance hierarchy is conserved among Brassicaceae. Here, we identified sRNAs and their target sites that can explain the linear dominance hierarchy of Arabidopsis lyrata, a species closely related to A. halleri. We tested the model that we established in Brassica to explain the linear dominance hierarchy in A. lyrata. Our results suggest that the dominance hierarchy of A. lyrata is also controlled by a homology-based interaction between sRNAs and their targets.     

Fluorescent whitening agents in Tokyo Bay and adjacent rivers: their application as anthropogenic molecular markers in coastal environments

Two kinds of stilbene-type fluorescent whitening agents (i.e., DSBP and DAS1), minor components of laundry detergents, were analyzed in surface waters of Tokyo Bay and adjacent rivers and in sewage effluents to examine their usefulness as molecular markers in the marine environment. Sensitive determination using HPLC (high performance liquid chromatography) with fluorescence detection with postcolumn UV radiator was employed. DSBP and DAS1 were found in Tokyo rivers at concentrations of a few microg/L and approximately 1 microg/L, respectively. DSBP and DAS1 were widely distributed in Tokyo Bay waters at concentrations in the range of 0.019-0.264 microg/L and 0.021-0.127 microg/L, respectively. Comparison of these concentrations with those in sewage effluents (DSBP: 8 microg/L and DAS1: 2.5 microg/L on average) yielded sewage dilutions in Tokyo Bay on the order of 10(2). FWAs-salinity diagram in the Tamagawa Estuary showed fairly conservative behaviors of the FWAs with approximately 20% and approximately 10% removal of DSBP and DAS1, respectively. This is thought to be caused by photodegradation. The persistent nature of FWAs and their widespread distribution in coastal environments demonstrates the utility of FWAs in tracing the behavior of water from rivers and sewage outfalls. The DSBP/DAS1 ratio showed a decreasing trend from sewage effluents, to rivers, to Tokyo Bay, indicating selective photodegradation of DSBP. The DSBP/DAS1 ratio is proposed as an index of the degree of photodegradation and residence time and freshness of water mass in coastal environments.     

Characterization and CO2 sorption properties of poly[methacryloxypropylheptacyclopentyl‐T8‐silsesquioxane‐ co‐3‐methacryloxypropyltris(trimethylsiloxy)silane]

Poly[methacryloxypropylheptacyclopentyl‐T8‐silsesquioxane (MAPOSS)‐co‐3‐methacryloxypropyltris(trimethylsiloxy)silane (SiMA)] was synthesized through free radical polymerization. The physical and carbon dioxide (CO₂) sorption properties of the copolymer membranes were investigated in terms of the MAPOSS content. As the MAPOSS content increases, the membrane density increased, suggesting a decrease in the fractional free volume. In addition, the thermal stability was improved with increasing the MAPOSS content. These are because of the polyhedraloligomericilsesquioxane (POSS) units that restrict the high mobility of poly(SiMA) segments. The glass transition temperature, T_g of the copolymers was single T_g based on the differential scanning calorimetry, suggesting that the copolymers were random and not phase separation. Based on the CO₂ sorption measurement, the POSS units play a role in reducing Henry's dissolution by suppressing the mobility of the poly(SiMA) component, while POSS units increase the nonequilibrium excess free volume, which contributes to the Langmuir dissolution. Based on these results, the introduction of MAPOSS unit is one of the effective ways to improved the thermal stability and CO₂ sorption property due to the enhancement of the polymer rigidity. © 2013 Wiley Periodicals, Inc. J. Appl. Polym. Sci., 2013