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991.
SNMP-1 (sensory neuron membrane protein 1) is an olfactory-specific membrane-bound protein which is homologous with the CD36 receptor family. Previous light level immunocytochemical studies suggested that SNMP-1 was localized in the dendrites and distal cell body of sex-pheromone-specific olfactory receptor neurons (ORN); these studies further suggested SNMP-1 was expressed in only one of two to three neurons in male-specific pheromone-sensitive trichoid sensilla. To better understand the expression and localization of SNMP-1, an immunocytochemical study was performed using electron microscopy to visualize the distribution of SNMP-1 among the neurons of several classes of olfactory sensilla of both male and female antennae of the silkmoth Antheraea polyphemus. SNMP-1 antigenicity was primarily restricted to the receptive dendritic membranes of ORNs of all sensilla types examined and was observed in cytosolic granules, but not plasma membranes, of the cell soma. Mean labeling densities ranged from 1 to 16 gold particles per micrometer of dendrite circumference; dendrites of trichoid and intermediate sensilla showed significantly higher labeling densities than those of basiconic sensilla. Larger dendrites of trichoid sensilla showed significantly higher mean labeling densities (13-16/micron) than smaller diameter dendrites (3-7/micron). Immunofluorescence studies using baculovirus expressed SNMP-1 and multiphoton photon laser scanning microscopy (MPLSM) indicated that rSNMP-1, which was post-translationally processed to the in vivo molecular weight, was inserted into the plasma membrane in a topography presenting extracellular epitopes. These studies suggest SNMP-1 is a common feature of the ORNs, is asymmetrically expressed among functionally distinct neurons, and possesses a topography which permits interaction with components of the extracellular sensillum lymph. 相似文献
992.
Disruption of a Global Regulatory Gene to Enhance Central Carbon Flux into Phenylalanine Biosynthesis in Escherichia coli 总被引:5,自引:0,他引:5
Genetic engineering of microbes for commercial metabolite production traditionally has sought to alter the levels and/or intrinsic
activities of key enzymes in relevant biosynthetic pathway(s). Microorganisms exploit similar strategies for flux control,
but also coordinate flux through sets of related pathways by using global regulatory circuits. We have engineered a global
regulatory system of Escherichia coli, Csr (carbon storage regulator), to increase precursor for aromatic amino acid biosynthesis. Disruption of csrA increases gluconeogenesis, decreases glycolysis, and thus elevates phosphoenolpyruvate, a limiting precursor of aromatics.
A strain in which the aromatic (shikimate) pathway had been optimized produced twofold more phenylalanine when csrA was disrupted. Overexpression of tktA (transketolase) to increase the other precursor, erythrose-4-phosphate, yielded ∼1.4-fold enhancement, while both changes
were additive. These effects of csrA were not mediated by increasing the regulatory enzymes of phenylalanine biosynthesis. This study introduces the concept of
“global metabolic engineering” for second-generation strain improvement.
Received: 25 October 2000 / Accepted: 8 December 2000 相似文献
993.
The purpose of this paper was to study the effect of the isopropyl myristic acid ester (IPM) on the physicochemical characteristics
of etoposide-loaded poly(lactic-co-glycolic acid) (PLGA) microspheres-specifically, the effects on the size and drug loading
of the microspheres, the polymer matrix and surface morphology, and the release of etoposide from the microspheres. The experiment
was structured to examine 2 IPM concentrations (25% and 50%) and 1 control (no IPM) at 2 different etoposide-loading percentages
(10% and 5%). The microspheres were prepared using a single-emulsion solvent-extraction procedure. Samples from each batch
of microspheres were then analyzed for size distribution. drug-loading efficiency, surface characteristics, in vitro release,
and in vitro microsphere degradation. The incorporation of 50% IPM significantly increased (P<05) the size of the microspheres when compared with the control and 25% IPM microspheres. However, incorporation of 25% or
50% IPM did not change (P>.05) the drug-loading efficiency in comparison with the microspheres prepared without IPM. The microspheres containing 50%
IPM were shown to significantly increase (P<.05) the release of etoposide from the microspheres at both etoposide concentrations. The microspheres prepared incorporating
25% IPM and 5% etoposide increased the in vitro release (P<.05) in comparison with the microspheres prepared without IPM. The 5% etoposide-PLGA microspheres showed a smooth, nonporous
surface that changed to a dimpled. nonporous surface after addition of 25% IPM. During the in vitro degradation study, the
IPM-containing microspheres slowly became porous but retained their structural integrity throughout the experiment. 相似文献
994.
Reticuloendothelial system (RES) particulate uptake (PU) of vascular debris influences survival from extreme hyperthermia. Little is known of the effect of extreme hyperthermia, unrelated to fever, on RES PU shortly after reaching a maximum core temperature (T(c)). Relative to normothermic rats (T(c)=38.0 degrees C), rats at T(c)=42.6 degrees C had significantly higher, while T(c)=42.0 degrees C rats had significantly lower total RES tissue (lung, liver, spleen) PU of fluorescent microspheres (1 μ), when compared to rats at T(c)=42.6 or 38.0 degrees C. These findings suggest at T(c)=42.6 degrees C, rats were not actively thermoregulating. As such, more blood remained in the core than in the periphery, which resulted in greater core RES tissue PU. In contrast, to reduce or control core heat, rats at T(c)=42.0 degrees or 38.0 degrees C directed more blood to the periphery, which reduced core RES tissue PU. Blood flow patterns as directed by the state or degree of active thermoregulation is likely an influence of hyperthermia on RES PU. 相似文献
995.
Joshua T. Ackerman Matthew C. Kondratieff Scott A. Matern Joseph J. Cech 《Environmental Biology of Fishes》2000,58(1):33-43
We used ultrasonic telemetry to determine the movement directions and movement rates of leopard sharks, Triakis semifasciata, in Tomales Bay, California. To analyze tide and time of day effects, we surgically implanted transmitters in the peritoneal cavities of one male and five female leopard sharks, which we located during summer for three to five sampling sessions lasting 12 to 24h each. All leopard sharks showed strong movement direction patterns with tide. During incoming tides, sharks moved significantly (p<0.0001) towards the inner bay, apparently to exploit the extensive inner bay muddy littoral zones' food resources. On outgoing tides, sharks showed significant (p<0.0001) movements towards the outer bay. During high tide, there was no discernible pattern to their movements (p=0.092). Shark movement rates were significantly (p<0.0001) greater during dark periods (mean±SE: 10.5±1.0m min–1), compared with fully lighted ones (6.7±0.5m min–1). Movement rates of longer sharks tended to be greater than those of shorter ones (range means±SE: 5.8±0.6m min–1 for the 91cm shark, to 12.8±1.6m min–1 for the 119cm shark), but the leopard sharks' overall mean movement rate (8.1±0.5m min–1) was slower than other (more pelagic) sharks. 相似文献
996.
To study the evolution of mtDNA and the intergeneric relationships of New World Jays (Aves: Corvidae), we sequenced the entire
mitochondrial DNA control region (CR) from 21 species representing all genera of New World jays, an Old World jay, crows,
and a magpie. Using maximum likelihood methods, we found that both the transition/transversion ratio (κ) and among site rate
variation (α) were higher in flanking domains I and II than in the conserved central domain and that the frequency of indels
was highest in domain II. Estimates of κ and α were much more influenced by the density of taxon sampling than by alternative
optimal tree topologies. We implemented a successive approximation method incorporating these parameters into phylogenetic
analysis. In addition we compared our study in detail to a previous study using cytochrome b and morphology to examine the effect of taxon sampling, evolutionary rates of genes, and combined data on tree resolution.
We found that the particular weighting scheme used had no effect on tree topology and little effect on tree robustness. Taxon
sampling had a significant effect on tree robustness but little effect on the topology of the best tree. The CR data set differed
nonsignificantly from the tree derived from the cytochrome b/morphological data set primarily in the placement of the genus Gymnorhinus, which is near the base of the CR tree. However, contrary to conventional taxonomy, the CR data set suggested that blue and
black jays (Cyanocorax sensu lato) might be paraphyletic and that the brown jay Psilorhinus (=Cyanocorax) morio is the sister group to magpie jays (Calocitta), a phylogenetic hypothesis that is likely as parsimonious with regard to nonmolecular characters as monophyly of Cyanocorax. The CR tree also suggests that the common ancestor of NWJs was likely a cooperative breeder. Consistent with recent systematic
theory, our data suggest that DNA sequences with high substitution rates such as the CR may nonetheless be useful in reconstructing
relatively deep phylogenetic nodes in avian groups.
Received: 10 November 1999 / Accepted: 16 March 2000 相似文献
997.
Matthew N.J. Seaman J. Michael McCaffery Scott D. Emr 《The Journal of cell biology》1998,142(3):665-681
We have recently characterized three yeast gene products (Vps35p, Vps29p, and Vps30p) as candidate components of the sorting machinery required for the endosome-to-Golgi retrieval of the vacuolar protein sorting receptor Vps10p (Seaman, M.N.J., E.G. Marcusson, J.-L. Cereghino, and S.D. Emr. 1997. J. Cell Biol. 137:79–92). By genetic and biochemical means we now show that Vps35p and Vps29p interact and form part of a multimeric membrane-associated complex that also contains Vps26p, Vps17p, and Vps5p. This complex, designated here as the retromer complex, assembles from two distinct subcomplexes comprising (a) Vps35p, Vps29p, and Vps26p; and (b) Vps5p and Vps17p. Density gradient fractionation of Golgi/endosomal/vesicular membranes reveals that Vps35p cofractionates with Vps5p/Vps17p in a vesicle-enriched dense membrane fraction. Furthermore, gel filtration analysis indicates that Vps35p and Vps5p are present on a population of vesicles and tubules slightly larger than COPI/coatomer-coated vesicles. We also show by immunogold EM that Vps5p is localized to discrete regions at the rims of the prevacuolar endosome where vesicles appear to be budding. Size fractionation of cytosolic and recombinant Vps5p reveals that Vps5p can self-assemble in vitro, suggesting that Vps5p may provide the mechanical impetus to drive vesicle formation. Based on these findings we propose a model in which Vps35p/Vps29p/Vps26p function to select cargo for retrieval, and Vps5p/Vps17p assemble onto the membrane to promote vesicle formation. Conservation of the yeast retromer complex components in higher eukaryotes suggests an important general role for this complex in endosome-to-Golgi retrieval. 相似文献
998.
Ratan V. Bhat Thomas M. Engber James P. Finn Elizabeth J. Koury Patricia C. Contreras Matthew S. Miller Craig A. Dionne Kevin M. Walton 《Journal of neurochemistry》1998,70(2):558-571
Abstract: In vitro studies indicate that p42/p44MAPK phosphorylate both nuclear and cytoplasmic proteins. However, the functional targets of p42/p44MAPK activation in vivo remain unclear. To address this question, we localized activated p42/p44MAPK in hippocampus and cortex and determined their signaling effects after electroconvulsive shock treatment (ECT) in rats. Phosphorylated p42/p44MAPK content increased in the cytoplasm of hippocampal neurons in response to ECT. Consistent with this cytoplasmic localization, inhibition of ECT-induced p42/p44MAPK activation by the extracellular signal-regulated kinase kinase inhibitor PD098059 blocked phosphorylation of the cytoplasmic protein microtubule-associated protein 2c (MAP2c), but failed to inhibit the induction of the nuclear protein c-Fos in response to ECT. In contrast to hippocampal neurons, cortical neurons exhibited an increase in amount of phosphorylated p42/p44MAPK in both the nucleus and cytoplasm after ECT. Accordingly, PD098059 blocked the induction of Fos-like immunoreactivity in the nuclei of cortical neurons as well as MAP2c phosphorylation in the cytoplasm. Our data indicate that both nuclear and cytoplasmic substrates can be activated by p42/p44MAPK in vivo. However, the functional targets of p42/p44MAPK signaling depend on the precise location of p42/p44MAPK within different subcellular compartments of brain regions. These results indicate unique functional pathways of p42/p44MAPK -mediated signal transduction within different brain regions in vivo. 相似文献
999.
Matthew Kennedy Lian Yu Maria João Lima Carla S. Ascenso Christopher Czaja Isabel Moura Jose J. G. Moura F. Rusnak 《Journal of biological inorganic chemistry》1998,3(6):643-649
Desulforedoxin and the N-terminus of desulfoferrodoxin share a 36 amino acid domain containing a (Cys-S)4 metal binding site. Recombinant forms of desulforedoxin, an N-terminal fragment of desulfoferrodoxin, and two desulforedoxin
mutant proteins were reconstituted with Fe3+, Cd2+, and Zn2+ and relative metal ion affinities assessed by proton titrations. Protons compete with metal for protein ligands, a process
that can be followed by monitoring the optical spectrum of the metal-protein complex as a function of pH. For all polypeptides,
Fe3+ bound with the highest affinity, whereas the affinity of Zn2+ was greater than Cd2+ in desulforedoxin and the N-terminal fragment of desulfoferrodoxin, but this order was reversed in desulforedoxin mutant
proteins. Metal binding in both mutants was significantly impaired. Furthermore, the Fe3+ complex of both mutants underwent a time-dependent bleaching process which coincided with increased reactivity of cysteine
residues to Ellman's reagent and concomitant metal dissociation. It is hypothesized that this results from an autoredox reaction
in which Fe3+ is reduced to Fe2+ with attendant oxidation of ligand thiols.
Received: 17 June 1998 / Accepted: 3 September 1998 相似文献
1000.