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91.
Sánchez-Sánchez F Aroca-Aguilar JD Segura I Ramírez-Castillejo C Riese HH Coca-Prados M Escribano J 《Journal of biotechnology》2008,134(1-2):193-201
Pigment epithelium-derived factor (PEDF) combines neurotrophic, neuroprotective, anti-angiogenic, anti-tumor and neural stem cell self-renewal properties in a single molecule, making this protein a valuable potential therapeutic agent. We herein analyzed the expression of human recombinant full-length PEDF, and its N- and C-terminal regions (amino acids 1-243 and 195-418, respectively) in three mammalian cell lines (HEK-293T, COS-1, and 26HCMsv), and in the yeast Pichia pastoris. The highest production of recombinant PEDF was achieved in P. pastoris which secreted approximately 30 microg of full-length rPEDF, and 47 microg of C-terminal/ml of culture medium. Full-length rPEDF was purified by one-step Ni-chelating high-performance liquid chromatography, recovering almost 70% of secreted rPEDF with a purity of 98.6%. The C-terminal region of PEDF was isolated by low-pressure liquid chromatography, recovering around 4% of the recombinant molecule with a purity of 98%. The N-terminal region of PEDF was not secreted by any expression system assayed. The two isolated recombinant PEDF polypeptides inhibited in vitro endothelial cell migration, and full-length rPEDF also increased cerebellar granule cell survival, thus demonstrating their biological activity. These polypeptides can be used to investigate the therapeutic role of PEDF in cancer, neurodegenerative and ocular diseases, and stem cell-based therapies. 相似文献
92.
Perálvarez-Marín A Lórenz-Fonfría VA Simón-Vázquez R Gomariz M Meseguer I Querol E Padrós E 《Biophysical journal》2008,95(9):4384-4395
Proline residues play a fundamental and subtle role in the dynamics, structure, and function in many membrane proteins. Temperature derivative spectroscopy and differential scanning calorimetry have been used to determine the effect of proline substitution in the structural stability of the active site and transmembrane arrangement of bacteriorhodopsin. We have analyzed the Pro-to-Ala mutation for the helix-embedded prolines Pro50, Pro91, and Pro186 in the native membrane environment. This information has been complemented with the analysis of the respective crystallographic structures by the FoldX force field. Differential scanning calorimetry allowed us to determine distorted membrane arrangement for P50A and P186A. The protein stability was severely affected for P186A and P91A. In the case of Pro91, a single point mutation is capable of strongly slowing down the conformational diffusion along the denaturation coordinate, becoming a barrier-free downhill process above 371 K. Temperature derivative spectroscopy, applied for first time to study thermal stability of proteins, has been used to monitor the stability of the active site of bacteriorhodopsin. The mutation of Pro91 and Pro186 showed the most striking effects on the retinal binding pocket. These residues are the Pro in closer contact to the active site (activation energies for retinal release of 60.1 and 76.8 kcal/mol, respectively, compared to 115.8 kcal/mol for WT). FoldX analysis of the protein crystal structures indicates that the Pro-to-Ala mutations have both local and long-range effects on the structural stability of residues involved in the architecture of the protein and the active site and in the proton pumping function. Thus, this study provides a complete overview of the substitution effect of helix-embedded prolines in the thermodynamic and dynamic stability of a membrane protein, also related to its structure and function. 相似文献
93.
Itziar Alba Marina Torreblanca Manuel Sánchez María Colom Inmaculada Meseguer 《Extremophiles : life under extreme conditions》2001,5(3):169-175
An organized structure, the fibrocrystalline body (FB), has been isolated from the archaeon Halobacterium salinarum. The structure is also present in, and can be isolated from, other extreme halophilic archaea. FB is present in the cytoplasm during the exponential growth and early stationary phases. This structure is affected by vincristine, an antitumoral drug, which targets tubulin. The drug causes fragmentation of the FB, changes in the cell shape, and growth inhibition. Taken together, these results point toward an important role in the life of the cell for this highly organized structure. 相似文献
94.
Juan M. Ruiz Rosa M. Rivero Inmaculada López-Cantarero Luis Romero 《Plant Growth Regulation》2003,41(2):173-177
Given the essential role played by phenol metabolism in many resistance responses to different types of stress, the aim of the present work was to determine how different application rates of calcium may influence this metabolic process. Increased calcium in the nutrient solution in which tobacco plants were grown considerably reduced the foliar concentration of phenolic compounds. Calcium clearly exerted a positive influence on the activities of enzymes (phenylalanine ammonia-lyase, polyphenol oxidase and peroxidase) involved in the metabolism of the phenolics. High dosages of calcium (5 mM) promoted more oxidation than synthesis of these compounds, thus explaining the lower concentration of the phenolics. 相似文献
95.
96.
J.H. Brock Inmaculada Esparza A.C. Logie 《Biochimica et Biophysica Acta (BBA)/General Subjects》1984,797(1):105-111
Mouse peritoneal macrophages were allowed to ingest 59Fe, 125I-labelled transferrin-antitransferrin immune complexes, and the release of 59Fe and degraded transferrin was studied. Some iron was released as ferritin, but a major portion was bound by bovine transferrin present in the culture medium, which contained fetal calf serum. If the medium was saturated with iron prior to incubation with the cells, little of the released iron was then bound by transferrin but appeared either as a high molecular weight fraction or, if nitrilotriacetate was present in the medium, some also appeared as a low molecular weight fraction. The release of non-ferritin iron was biphasic, the early, rapid phase being more prolonged with resident cells than with stimulated cells. The rate of release in the late phase did not differ significantly between resident and stimulated cells. Incubation at 0°C completely suppressed the release of degraded transferrin, but iron release continued at about 30% of the rate seen in control cultures at 37°C. A model for the intracellular handling of ingested iron is proposed to take account of the different release patterns of resident and stimulated macrophages. 相似文献
97.
Rafael Tormo Molina Adolfo Muñoz Rodríguez Inmaculada Silva Palacios 《Aerobiologia》1996,12(1):161-166
Two years of data from four longitudinal traverses along each day's slide prepared from a continuously running Burkard sporetrap have been analyzed statistically. Using the Friedman test, a statistically significant difference was found between the four traverses, with a greater than 7% loss of pollen grains in the two outer traverses in relation to the inner. Four slides were then selected for more detailed analysis, using 18 longitudinal traverses with a 1-mm separation from the upper to the lower edge of the Melinex tape. There was found to be a progressive decline from the centre to the outside, and more than 4% of pollen grains were found outside the typical 14 mm width of the impaction orifice. There was no correlation between pollen grain size and the decline in counts from the centre to the outside. For the complete data set, there was a general rise in the diversity of pollen types with increasing sample counts, but above about 1000 pollen grains per sample there were no more than 27 pollen types found, often even fewer. A discussion is presented of whether four traverses really should be a fixing sample size, or whether it might be better to fix the total pollen count beginning with a traverse in the middle of the slide and ending with a variable number of traverses when that count is reached. 相似文献
98.
99.
Alejandro Monroy-Colín Inmaculada Silva-Palacios Rafael Tormo-Molina José María Maya-Manzano Santiago Fernández Rodríguez Ángela Gonzalo-Garijo 《Aerobiologia》2018,34(3):269-283
Narrow-leafed ash (Fraxinus angustifolia) is a common polygamous tree growing on the banks of rivers in the western Mediterranean region. Pollination occurs during winter, and the tree’s pollen is among the most abundant during that season. This work aims to relate the phenology of pollen shedding, source tree distribution, meteorology and airborne pollen occurrence for the species. Aerobiological sampling was conducted in Badajoz (south-western Spain) using a Hirst volumetric sampler over 24 years (1993–2016). Trees were geo-localized in a circle 500 m in diameter surrounding the pollen sampler. During the last two periods, pollination phenology was studied in 10 specimens, five in the surroundings of the pollen station and five 3 km apart, at a frequency of 3–4 days on average. Moreover, a detailed analysis of pollen occurrence was conducted for these two periods. Daily data for the whole period and hourly data over the last 2 years were used, including pollen monitoring and meteorology. A comparison was made between pollen occurrence and source distribution. The main pollen season lasted on average 53 (28–75) days. Average values were less than 10 grains m?3, except for two periods of 23–24 grains m?3. Daily data and hourly data correlation with meteorology showed different signs in correlation analysis. Hourly analysis showed that the maximum concentration occurred just after noon. Most pollen was recorded at an average temperature of 9 °C. Analysis of pollen sources and pollen occurrence showed a close relationship between predominant wind directions and tree distribution. Peaks of phenology were not coincident with pollen peaks. No trends in pollination were found. Non-homogeneous distribution of pollen sources for Fraxinus angustifolia provided a suitable tool to demonstrate that wind direction plays a relevant role when aerobiological data are interpreted according to source distribution. A limitation in phenology analysis and aerobiological data was noted in the narrow-leafed ash species. 相似文献
100.
Inmaculada Castro José Espinosa José E. Granados Francisco J. Cano-Manuel Paulino Fandos Arián Ráez-Bravo Jorge R. López-Olvera Ramón C. Soriguer Jesús M. Pérez 《Experimental & applied acarology》2018,75(1):41-53
The plant stress hypothesis posits that a herbivore’s reproductive success increases when it feeds on stressed plants, while the plant vigor hypothesis predicts that a herbivore preferentially feeds on more vigorous plants. We examined these opposing hypotheses by growing spider mites (Tetranychus urticae) on the leaves of stressed and healthy (vigorous) cucumber plants. Host plants were grown under controlled conditions at low, moderate, and high concentrations of NaCl (to induce salinity stress), at low, moderate, and high fertilizer concentrations (to support growth), and without these additions (control). The effects of these treatments were evaluated by measuring fresh and dry plant biomass, carotenoid and chlorophyll content, antioxidant enzyme activity, and concentrations of PO43?, K+, and Na+ in plant tissues. The addition of low concentrations of fertilizer increased dry mass, protein, and carotenoid content relative to controls, suggesting a beneficial effect on plants. The highest NaCl treatment (2560 mg L?1) resulted in increased Na+ and protein content relative to control plants, as well as reduced PO43?, K+, and chlorophyll levels and reduced catalase and ascorbate peroxidase enzyme activity levels. Analysis of life table data of T. urticae mites raised on leaves from the aforementioned plant groups showed the intrinsic rate of increase (r) for mites was 0.167 day?1 in control specimens, 0.125 day?1 for mites reared on plants treated with a moderate concentration of fertilizer (10 mL L?1), and was highest (0.241 day?1) on plants grown under moderate salinity conditions (1920 mg L?1 NaCl). Reproductive success of T. urticae did not differ on plants watered with a moderate concentration of NaCl or a high concentration of fertilizer. The moderately-stressed plants formed a favorable environment for the development and reproduction of spider mites, supporting the plant stress hypothesis. 相似文献