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971.
Evolutionary studies often estimate fitness components with the aim to make predictions about the outcome of selection. Depending
on the system and the question, different fitness components are used, but their usefulness for predicting the outcome of
selection is rarely tested. Here we estimate host fitness components in different ways with the aim to test how well they
agree with each other and how well they predict host fitness at the population level in the presence of the parasite. We use
a Daphnia magna-microparasite system to study the competitive ability of host clones in the absence and presence of the parasite, the infection
intensity of the parasite in individuals of twelve host clones (an estimate of both host resistance and parasite reproductive
success), and parasite persistence in small host populations (an estimate of R
0 of the parasite). Analysis of host competitive ability and parasite persistence reveals strong host genotype effects, while
none are found for infection intensity. Host competitive ability further shows a genotype-specific change upon infection,
which is correlated with the relative persistence of the parasite in the competing hosts. Hosts in which the parasite persists
better suffer a competitive disadvantage in the parasite’s presence. This suggests that in this system, parasite-mediated
selection can be predicted by parasite persistence, but not by parasite infection intensity. 相似文献
972.
Mechanical single molecule techniques offer exciting possibilities for investigating protein folding and stability in native environments at sub-nanometer resolutions. Compatible solutes show osmotic activity which even at molar concentrations do not interfere with cell metabolism. They are known to protect proteins against external stress like temperature, high salt concentrations and dehydrating conditions. We studied the impact of the compatible solute ectoine (1M) on membrane proteins by analyzing the mechanical properties of Bacteriorhodopsin (BR) in its presence and absence by single molecule force spectroscopy. The unfolding experiments on BR revealed that ectoine decreases the persistence length of its polypeptide chain thereby increasing its tendency to coil up. In addition, we found higher unfolding forces indicating strengthening of those intra molecular interactions which are crucial for stability. This shows that force spectroscopy is well suited to study the effect of compatible solutes to stabilize membrane proteins against unfolding. In addition, it may lead to a better understanding of their detailed mechanism of action. 相似文献
973.
Cell wall-degrading enzymes of Venturia inaequalis are supposed to be fungal virulence factors whereas phenolic compounds of the host plant may be involved in defence. Since
phenolic structures are predestined for an interaction with proteins we studied the effects on enzymes and proteins in course
of in vitro culture and with preparations from culture filtrates and mycelia, respectively. The native compounds epicatechin,
catechin, phloridzin, chlorogenic acid, caffeic acid, p-coumaric acid and phloridzin tested under non-oxidizing conditions had no or weak effects on enzyme activities. A significant inhibition of pectinase was only detected with the highest concentrations of procyanidins and phloretin. Aerobe
conditions resulted in a fast oxidation of most phenolics which was enhanced by fungal phenoloxidases. Generally, no inhibition
of fungal growth occurred in vitro but distinct irreversible effects on proteins and enzymes were detected with oxidized phenolics
in course of in vitro-cultures as well as with the corresponding preparations. Efficacy of inhibitory activity in in vitro-cultures
depended on media, culture technique and time course. Direct treatment of enzyme preparations with the oxidized phenolics
resulted in a distinct inhibition of cellulolytic and especially pectinolytic activity. Apart from cellulase pattern altered
by phenolics, in vitro-culture zymograms revealed a non specific reduction of enzymatic activities, whereas action on total
culture filtrate proteins resulted in specific effects due to phenolic compounds and incubation time. An attempt was made
to characterize the oxidation products of epicatechin. Chromatographic fractionation revealed a non-resolvable complex of
inhibitory compounds which were not consistent with the typical yellow oxidation products. 相似文献
974.
Dieter Deryckere Tom Eeckhaut Johan Van Huylenbroeck Erik Van Bockstaele 《Plant cell reports》2012,31(12):2261-2269
Key message
A standard method has been developed with which we are able to fully regenerate protoplasts of different Cichorium species. For the first time, endive protoplasts have been regenerated into plantlets.Abstract
Protoplast regeneration is essential for somatic hybridizations. In this study, a standard method for plantlet regeneration from Cichorium protoplasts was developed. We evaluated the effect of the low melting point agarose (LMPA) bead technique on the regeneration capacity of protoplasts of seven C. intybus and four C. endivia genotypes. The LMPA bead technique was more efficient than culture in liquid or solid medium and allowed us to obtain plating efficiencies up to 4.9?% in C. intybus genotypes and efficiencies of up to 0.7?% in C. endivia genotypes. Moreover, the LMPA bead technique offers great advantages over liquid and solid culture systems: the media can be readily refreshed, protoplasts can be monitored separately, and microcalli can easily be removed from the beads. This increased efficiency was observed for all of the 11 Cichorium genotypes tested. Shoot formation was induced more efficiently when using 0.5?mg?l?1 indole-3-acetic acid-enriched medium (up to 87.5?% of the protoplast-derived calli started shoot development) compared to 1-naphthaleneacetic acid-enriched medium. The LMPA bead technique optimized in this study enabled for the first time the full plantlet regeneration from protoplasts of C. endivia genotypes and increased the protoplast regenerating ability in other Cichorium species. This fine-tuned LMPA bead technique can therefore be applied for protoplast regeneration after protoplast fusions of the genus Cichorium. 相似文献975.
976.
Nimishakavi S Besprozvannaya M Raymond WW Craik CS Gruenert DC Caughey GH 《American journal of physiology. Lung cellular and molecular physiology》2012,303(2):L97-106
Prostasin is a membrane-anchored protease expressed in airway epithelium, where it stimulates salt and water uptake by cleaving the epithelial Na(+) channel (ENaC). Prostasin is activated by another transmembrane tryptic protease, matriptase. Because ENaC-mediated dehydration contributes to cystic fibrosis (CF), prostasin and matriptase are potential therapeutic targets, but their catalytic competence on airway epithelial surfaces has been unclear. Seeking tools for exploring sites and modulation of activity, we used recombinant prostasin and matriptase to identify substrate t-butyloxycarbonyl-l-Gln-Ala-Arg-4-nitroanilide (QAR-4NA), which allowed direct assay of proteases in living cells. Comparisons of bronchial epithelial cells (CFBE41o-) with and without functioning cystic fibrosis transmembrane conductance regulator (CFTR) revealed similar levels of apical and basolateral aprotinin-inhibitable activity. Although recombinant matriptase was more active than prostasin in hydrolyzing QAR-4NA, cell surface activity resisted matriptase-selective inhibition, suggesting that prostasin dominates. Surface biotinylation revealed similar expression of matriptase and prostasin in epithelial cells expressing wild-type vs. ΔF508-mutated CFTR. However, the ratio of mature to inactive proprostasin suggested surface enrichment of active enzyme. Although small amounts of matriptase and prostasin were shed spontaneously, prostasin anchored to the cell surface by glycosylphosphatidylinositol was the major contributor to observed QAR-4NA-hydrolyzing activity. For example, the apical surface of wild-type CFBE41o- epithelial cells express 22% of total, extractable, aprotinin-inhibitable, QAR-4NA-hydrolyzing activity and 16% of prostasin immunoreactivity. In conclusion, prostasin is present, mature and active on the apical surface of wild-type and CF bronchial epithelial cells, where it can be targeted for inhibition via the airway lumen. 相似文献
977.
Wan Y Jasik J Wang L Hao H Volkmann D Menzel D Mancuso S Baluška F Lin J 《The Plant cell》2012,24(2):551-565
Under blue light (BL) illumination, Arabidopsis thaliana roots grow away from the light source, showing a negative phototropic response. However, the mechanism of root phototropism is still unclear. Using a noninvasive microelectrode system, we showed that the BL sensor phototropin1 (phot1), the signal transducer NONPHOTOTROPIC HYPOCOTYL3 (NPH3), and the auxin efflux transporter PIN2 were essential for BL-induced auxin flux in the root apex transition zone. We also found that PIN2-green fluorescent protein (GFP) localized to vacuole-like compartments (VLCs) in dark-grown root epidermal and cortical cells, and phot1/NPH3 mediated a BL-initiated pathway that caused PIN2 redistribution to the plasma membrane. When dark-grown roots were exposed to brefeldin A (BFA), PIN2-GFP remained in VLCs in darkness, and BL caused PIN2-GFP disappearance from VLCs and induced PIN2-GFP-FM4-64 colocalization within enlarged compartments. In the nph3 mutant, both dark and BL BFA treatments caused the disappearance of PIN2-GFP from VLCs. However, in the phot1 mutant, PIN2-GFP remained within VLCs under both dark and BL BFA treatments, suggesting that phot1 and NPH3 play different roles in PIN2 localization. In conclusion, BL-induced root phototropism is based on the phot1/NPH3 signaling pathway, which stimulates the shootward auxin flux by modifying the subcellular targeting of PIN2 in the root apex transition zone. 相似文献
978.
Dutt S Kléber M Matasci M Sommer L Zimmermann DR 《The Journal of biological chemistry》2006,281(17):12123-12131
We previously showed the selective expression of the chondroitin sulfate proteoglycans versican V0 and V1 in barrier tissues that impede the migration of neural crest cells during embryonic trunk development (Landolt, R. M., Vaughan, L., Winterhalter, K. H., and Zimmermann, D. R. (1995) Development 212, 2303-2312). To test for an active involvement of these isoforms in the guidance process, we have now established protocols to isolate intact versican V0 and V1 in quantities sufficient for functional experiments. Using stripe choice assays, we demonstrate that pure preparations of either a mixture of versican V0/V1 or V1 alone strongly inhibit the migration of multipotent Sox10/p75NTR double-positive early neural crest stem cells on fibronectin by interfering with cell-substrate adhesion. We show that this inhibition is largely core glycoprotein-dependent, as the complete removal of the glycosaminoglycan chains has only a minor effect on the inhibitory capacity. Our findings support the notion that versican variants V0 and V1 act, possibly in concert with other inhibitory molecules such as aggrecan and ephrins, in directing the migratory streams of neural crest cells to their appropriate target tissues. 相似文献
979.
Leupold D Teuchner K Ehlert J Irrgang KD Renger G Lokstein H 《The Journal of biological chemistry》2006,281(35):25381-25387
Stepwise two-photon excited fluorescence (TPEF) spectra of the photosynthetic antenna complexes PCP, CP47, CP29, and light-harvesting complex II (LHC II) were measured. TPEF emitted from higher excited states of chlorophyll (Chl) a and b was elicited via consecutive absorption of two photons in the Chl a/b Qy range induced by tunable 100-fs laser pulses. Global analyses of the TPEF line shapes with a model function for monomeric Chl a in a proteinaceous environment allow distinction between contributions from monomeric Chls a and b, strongly excitonically coupled Chls a, and Chl a/b heterodimers/-oligomers. The analyses indicate that the longest wavelength-absorbing Chl species in the Qy region of LHC II is a Chl a homodimer with additional contributions from adjacent Chl b. Likewise, in CP47 a spectral form at approximately 680 nm (that is, however, not the red-most species) is also due to strongly coupled Chls a. In contrast to LHC II, the red-most Chl subband of CP29 is due to a monomeric Chl a. The two Chls b in CP29 exhibit marked differences: a Chl b absorbing at approximately 650 nm is not excitonically coupled to other Chls. Based on this finding, the refractive index of its microenvironment can be determined to be 1.48. The second Chl b in CP29 (absorbing at approximately 640 nm) is strongly coupled to Chl a. Implications of the findings with respect to excitation energy transfer pathways and rates are discussed. Moreover, the results will be related to most recent structural analyses. 相似文献
980.
Batra-Safferling R Abarca-Heidemann K Körschen HG Tziatzios C Stoldt M Budyak I Willbold D Schwalbe H Klein-Seetharaman J Kaupp UB 《The Journal of biological chemistry》2006,281(3):1449-1460
The outer segment of vertebrate photoreceptors is a specialized compartment that hosts all the signaling components required for visual transduction. Specific to rod photoreceptors is an unusual set of three glutamic acid-rich proteins (GARPs) as follows: two soluble forms, GARP1 and GARP2, and the N-terminal cytoplasmic domain (GARP' part) of the B1 subunit of the cyclic GMP-gated channel. GARPs have been shown to interact with proteins at the rim of the disc membrane. Here we characterized native GARP1 and GARP2 purified from bovine rod photoreceptors. Amino acid sequence analysis of GARPs revealed structural features typical of "natively unfolded" proteins. By using biophysical techniques, including size-exclusion chromatography, dynamic light scattering, NMR spectroscopy, and circular dichroism, we showed that GARPs indeed exhibit a large degree of intrinsic disorder. Analytical ultracentrifugation and chemical cross-linking showed that GARPs exist in a monomer/multimer equilibrium. The results suggested that the function of GARP proteins is linked to their structural disorder. They may provide flexible spacers or linkers tethering the cyclic GMP-gated channel in the plasma membrane to peripherin at the disc rim to produce a stack of rings of these protein complexes along the long axis of the outer segment. GARP proteins could then provide the environment needed for protein interactions in the rim region of discs. 相似文献