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811.
812.
Keating E Waring AJ Walther FJ Possmayer F Veldhuizen RA Petersen NO 《Biochimica et biophysica acta》2011,1808(3):614-621
Pulmonary surfactant is a complex lipid-protein mixture whose main function is to reduce the surface tension at the air-liquid interface of alveoli to minimize the work of breathing. The exact mechanism by which surfactant monolayers and multilayers are formed and how they lower surface tension to very low values during lateral compression remains uncertain. We used time-of-flight secondary ion mass spectrometry to study the lateral organization of lipids and peptide in surfactant preparations ranging in complexity. We show that we can successfully determine the location of phospholipids, cholesterol and a peptide in surfactant Langmuir-Blodgett films and we can determine the effect of cholesterol and peptide addition. A thorough understanding of the lateral organization of PS interfacial films will aid in our understanding of the role of each component as well as different lipid-lipid and lipid-protein interactions. This may further our understanding of pulmonary surfactant function. 相似文献
813.
Investigations of K+-occlusion by the phosphoenzyme of Na+,K+-ATPase from shark rectal gland and pig kidney by stopped-flow fluorimetry reveal major differences in the kinetics of the two enzymes. In the case of the pig enzyme, a single K+-occlusion step could be resolved with a rate constant of 342 (±26) s−1. However, in the case of the shark enzyme, two consecutive K+-occlusions were detected with rate constants of 391 (±19) s−1 and 48 (±2) s−1 at 24°C and pH 7.4. A conformational change of the phosphoenzyme associated with K+-occlusion is, thus, the major rate-determining step of the shark enzyme under saturating concentrations of all substrates, whereas for the pig enzyme the major rate-determining step under the same conditions is the E2 → E1 transition and its associated K+ deocclusion and release to the cytoplasm. The differences in rate constants of the K+ occlusion reactions of the two enzymes are paralleled by compensating changes to the rate constant for the E2 → E1 transition, which explains why the differences in the enzymes' kinetic behaviors have not previously been identified. 相似文献
814.
Madic J Vingadassalon N de Garam CP Marault M Scheutz F Brugère H Jamet E Auvray F 《Applied and environmental microbiology》2011,77(6):2035-2041
Shiga toxin (Stx)-producing Escherichia coli (STEC) strains are a diverse group of food-borne pathogens with various levels of virulence for humans. In this study, we describe the use of a combination of multiple real-time PCR assays for the screening of 400 raw-milk cheeses for the five main pathogenic STEC serotypes (O26:H11, O103:H2, O111:H8, O145:H28, and O157:H7). The prevalences of samples positive for stx, intimin-encoding gene (eae), and at least one of the five O group genetic markers were 29.8%, 37.3%, and 55.3%, respectively. The H2, H7, H8, H11, and H28 fliC alleles were highly prevalent and could not be used as reliable targets for screening. Combinations of stx, eae variants, and O genetic markers, which are typical of the five targeted STEC serotypes, were detected by real-time PCR in 6.5% of the cheeses (26 samples) and included stx-wzx(O26)-eae-β1 (4.8%; 19 samples), stx-wzx(O103)-eae-ε (1.3%; five samples), stx-ihp1(O145)-eae-γ1 (0.8%; three samples), and stx-rfbE(O157)-eae-γ1 (0.3%; one sample). Twenty-eight immunomagnetic separation (IMS) assays performed on samples positive for these combinations allowed the recovery of seven eaeβ1-positive STEC O26:H11 isolates, whereas no STEC O103:H2, O145:H28, or O157:H7 strains could be isolated. Three stx-negative and eaeβ1-positive E. coli O26:[H11] strains were also isolated from cheeses by IMS. Colony hybridization allowed us to recover STEC from stx-positive samples for 15 out of 45 assays performed, highlighting the difficulties encountered in STEC isolation from dairy products. The STEC O26:H11 isolates shared the same virulence genetic profile as enterohemorrhagic E. coli (EHEC) O26:H11, i.e., they carried the virulence-associated genes EHEC-hlyA, katP, and espP, as well as genomic O islands 71 and 122. Except for one strain, they all contained the stx1 variant only, which was reported to be less frequently associated with human cases than stx2. Pulsed-field gel electrophoresis (PFGE) analysis showed that they displayed high genetic diversity; none of them had patterns identical to those of human O26:H11 strains investigated here. 相似文献
815.
Karotki L Huiskonen JT Stefan CJ Ziółkowska NE Roth R Surma MA Krogan NJ Emr SD Heuser J Grünewald K Walther TC 《The Journal of cell biology》2011,195(5):889-902
Spatial organization of membranes into domains of distinct protein and lipid composition is a fundamental feature of biological systems. The plasma membrane is organized in such domains to efficiently orchestrate the many reactions occurring there simultaneously. Despite the almost universal presence of membrane domains, mechanisms of their formation are often unclear. Yeast cells feature prominent plasma membrane domain organization, which is at least partially mediated by eisosomes. Eisosomes are large protein complexes that are primarily composed of many subunits of two Bin-Amphiphysin-Rvs domain-containing proteins, Pil1 and Lsp1. In this paper, we show that these proteins self-assemble into higher-order structures and bind preferentially to phosphoinositide-containing membranes. Using a combination of electron microscopy approaches, we generate structural models of Pil1 and Lsp1 assemblies, which resemble eisosomes in cells. Our data suggest that the mechanism of membrane organization by eisosomes is mediated by self-assembly of its core components into a membrane-bound protein scaffold with lipid-binding specificity. 相似文献
816.
Flemming TM 《Behavioural processes》2011,86(3):316-322
Learning of the relational same/different (S/D) concept has been demonstrated to be largely dependent upon stimulus sets containing more than two items for pigeons and old-world monkeys. Stimulus arrays containing several images for use in same/different discrimination procures (e.g. 16 identical images vs. 16 nonidentical images) have been shown to facilitate and even be necessary for learning of relational concepts (
[Flemming et al., 2007],
[Wasserman et al., 2001] and [Young et al., 1997]). In the present study, we investigate the threshold at which a new world primate, the capuchin (Cebus apella) may be able to make such a discrimination. Utilizing a method of increasing entropy, rather than conventional procedures of decreasing entropy, we demonstrate unique evidence that capuchin monkeys are readily capable of making 2-item relational S/D conditional discriminations. In another experiment, we examine the supposed level of difficulty in making S/D discriminations by rhesus monkeys (Macaca mulatta). Whereas pigeons (Columba livia) and baboons (Papio papio) have shown marked difficulty simultaneously discriminating same from different arrays at all when composed of fewer than 8 items each, rhesus monkeys seem to understand that pairs of stimuli connote sameness and difference just the same (Flemming et al., 2007). With sustained accurate performance of 2-item S/D discriminations, both experienced and task-naïve rhesus monkeys appear quite certain in their conceptual knowledge of same and different. We conclude that learning of the same/different relational concept may be less dependent upon high levels of entropy contrast than originally hypothesized for nonhuman primates. 相似文献
817.
We describe YGFP, a slow bleaching green fluorescent protein (GFP) with unique spectral properties. YGFP is derived from an Escherichia coli codon-optimized synthetic gfp mutant 2 derivative. In addition to the GFP-mut 2 changes, it also carries S202F and T203I substitutions. YGFP can be used as a substitute for yellow fluorescent protein (YFP) in experiments in which two or more fluorescent proteins are fused to different cellular protein components, expanding the ability to study multiple labeled proteins in a cell at once. 相似文献
818.
Random coil chemical shifts are necessary for secondary chemical shift analysis, which is the main NMR method for identification
of secondary structure in proteins. One of the largest challenges in the determination of random coil chemical shifts is accounting
for the effect of neighboring residues. The contributions from the neighboring residues are typically removed by using neighbor
correction factors determined based on each residue’s effect on glycine chemical shifts. Due to its unusual conformational
freedom, glycine may be particularly unrepresentative for the remaining residue types. In this study, we use random coil peptides
containing glutamine instead of glycine to determine the random coil chemical shifts and the neighbor correction factors.
The resulting correction factors correlate to changes in the populations of the major wells in the Ramachandran plot, which
demonstrates that changes in the conformational ensemble are an important source of neighbor effects in disordered proteins.
Glutamine derived random coil chemical shifts and correction factors modestly improve our ability to predict 13C chemical shifts of intrinsically disordered proteins compared to existing datasets, and may thus improve the identification
of small populations of transient structure in disordered proteins. 相似文献
819.
820.
Alferso C. Abrahams Sayed M. Habib Amélie Dendooven Bruce L. Riser Jan Willem van der Veer Raechel J. Toorop Michiel G. H. Betjes Marianne C. Verhaar Christopher J. E. Watson Tri Q. Nguyen Walther H. Boer 《PloS one》2014,9(11)