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1.
Vibrio cholerae is autochthonous to various aquatic niches and is the etiological agent of the life-threatening diarrheal disease cholera. The persistence of V. cholerae in natural habitats is a crucial factor in the epidemiology of cholera. In contrast to the well-studied V. cholerae-chitin connection, scarce information is available about the factors employed by the bacteria for the interaction with collagens. Collagens might serve as biologically relevant substrates, because they are the most abundant protein constituents of metazoan tissues and V. cholerae has been identified in association with invertebrate and vertebrate marine animals, as well as in a benthic zone of the ocean where organic matter, including collagens, accumulates. Here, we describe the characterization of the V. cholerae putative collagenase, VchC, encoded by open reading frame VC1650 and belonging to the subfamily M9A peptidases. Our studies demonstrate that VchC is an extracellular collagenase degrading native type I collagen of fish and mammalian origin. Alteration of the predicted catalytic residues coordinating zinc ions completely abolished the protein enzymatic activity but did not affect the translocation of the protease by the type II secretion pathway into the extracellular milieu. We also show that the protease undergoes a maturation process with the aid of a secreted factor(s). Finally, we propose that V. cholerae is a collagenovorous bacterium, as it is able to utilize collagen as a sole nutrient source. This study initiates new lines of investigations aiming to uncover the structural and functional components of the V. cholerae collagen utilization program.  相似文献   
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Unequal absorption of photons between photosystems I and II, and between bundle-sheath and mesophyll cells, are likely to affect the efficiency of the CO2-concentrating mechanism in C4 plants. Under steady-state conditions, it is expected that the biochemical distribution of energy (ATP and NADPH) and photosynthetic metabolite concentrations will adjust to maintain the efficiency of C4 photosynthesis through the coordination of the C3 (Calvin-Benson-Bassham) and C4 (CO2 pump) cycles. However, under transient conditions, changes in light quality will likely alter the coordination of the C3 and C4 cycles, influencing rates of CO2 assimilation and decreasing the efficiency of the CO2-concentrating mechanism. To test these hypotheses, we measured leaf gas exchange, leaf discrimination, chlorophyll fluorescence, electrochromatic shift, photosynthetic metabolite pools, and chloroplast movement in maize (Zea mays) and Miscanthus × giganteus following transitional changes in light quality. In both species, the rate of net CO2 assimilation responded quickly to changes in light treatments, with lower rates of net CO2 assimilation under blue light compared with red, green, and blue light, red light, and green light. Under steady state, the efficiency of CO2-concentrating mechanisms was similar; however, transient changes affected the coordination of C3 and C4 cycles in M. giganteus but to a lesser extent in maize. The species differences in the ability to coordinate the activities of C3 and C4 cycles appear to be related to differences in the response of cyclic electron flux around photosystem I and potentially chloroplast rearrangement in response to changes in light quality.The CO2-concentrating mechanism in C4 plants reduces the carbon lost through the photorespiratory pathway by limiting the oxygenation of ribulose-1,5-bisphosphate (RuBP) by the enzyme Rubisco (Brown and Smith, 1972; Sage, 1999). Through the compartmentalization of the C4 cycle in the mesophyll cells and the C3 cycle in the bundle-sheath cells (Hatch and Slack, 1966), C4 plants suppress RuBP oxygenation by generating a high CO2 partial pressure around Rubisco (Furbank and Hatch, 1987). To maintain high photosynthetic rates and efficient light energy utilization, the metabolic flux through the C3 and C4 cycles must be coordinated. However, coordination of the C3 and C4 cycles is likely disrupted due to rapid changes in environmental conditions, particularly changes in light availability (Evans et al., 2007; Tazoe et al., 2008).Spatial and temporal variations in light environments, including both light quantity and quality, are expected to alter the coordination of the C3 and C4 cycles. For example, it has been suggested that the coordination of C3 and C4 cycles is altered by changes in light intensity (Henderson et al., 1992; Cousins et al., 2006; Tazoe et al., 2006, 2008; Kromdijk et al., 2008, 2010; Pengelly et al., 2010). However, more recent publications indicate that some of the proposed light sensitivity of the CO2-concentrating mechanisms in C4 plants can be attributed to oversimplifications of leaf models of carbon isotope discrimination (Δ13C), in particular, errors in estimates of bundle-sheath CO2 partial pressure and omissions of respiratory fractionation (Ubierna et al., 2011, 2013). Alternatively, there is little information on the effects of light quality on the coordination of C3 and C4 cycle activities and the subsequent impact on net rate of CO2 assimilation (Anet).In C3 plants, Anet is reduced under blue light compared with red or green light (Evans and Vogelmann, 2003; Loreto et al., 2009). This was attributed to differences in absorbance and wavelength-dependent differences in light penetration into leaves, where red and green light penetrate farther into leaves compared with blue light (Vogelmann and Evans, 2002; Evans and Vogelmann, 2003). Differences in light quality penetration into a leaf are likely to have profound impacts on C4 photosynthesis, because the C4 photosynthetic pathway requires the metabolic coordination of the mesophyll C4 cycle and the bundle-sheath C3 cycle. Indeed, Evans et al. (2007) observed a 50% reduction in the rate of CO2 assimilation in Flaveria bidentis under blue light relative to white light at a light intensity of 350 µmol quanta m−2 s−1. This was attributed to poor penetration of blue light into the bundle-sheath cells and subsequent insufficient production of ATP in the bundle-sheath cells to match the rates of mesophyll cell CO2 pumping (Evans et al., 2007). Recently, Sun et al. (2012) observed similar low rates of steady-state CO2 assimilation under blue light relative to red, green, and blue light (RGB), red light, and green light at a constant light intensity of 900 µmol quanta m−2 s−1.Because the light penetration into a leaf depends on light quality, with blue light penetrating the least, this potentially results in changes in the energy available for carboxylation reactions in the bundle-sheath (C3 cycle) and mesophyll (C4 cycle) cells. Changes in the balance of energy driving the C3 and C4 cycles can alter the efficiency of the CO2-concentrating mechanisms, often represented by leakiness (ϕ), the fraction of CO2 that is pumped into the bundle-sheath cells that subsequently leaks back out (Evans et al., 2007). Unfortunately, ϕ cannot be measured directly, but it can be estimated through the combined measured and modeled values of Δ13C (Farquhar, 1983). Using measurements of Δ13C, it has been demonstrated that under steady-state conditions, changes in light quality do not affect ϕ (Sun et al., 2012); however, it remains unknown if ϕ is also constant during the transitions between different light qualities. In fact, sudden changes of light quality could temporally alter the coordination of the C3 and C4 cycles.To understand the effects of light quality on C4 photosynthesis and the coordination of the activities of C3 and C4 cycles, we measured transitional changes in leaf gas exchange and Δ13C under RGB and broad-spectrum red, green, and blue light in the NADP-malic enzyme C4 plants maize (Zea mays) and Miscanthus × giganteus. Leaf gas exchange and Δ13C measurements were used to estimate ϕ using the complete model of C4 leaf Δ13C (Farquhar, 1983; Farquhar and Cernusak, 2012). Additionally, we measured photosynthetic metabolite pools, Rubisco activation state, chloroplast movement, and rates of linear versus cyclic electron flow during rapid transitions from red to blue light and blue to red light. We hypothesized that the limited penetration of blue light into the leaf would result in insufficient production of ATP in the bundle-sheath cells to match the rate of mesophyll cell CO2 pumping. We predicted that rapid changes in light quality would affect the coordination of the C3 and C4 cycles and cause an increase in ϕ, but this would equilibrate as leaf metabolism reached a new steady-state condition.  相似文献   
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Autophagy, the process by which proteins or organelles are engulfed by autophagosomes and delivered for vacuolar/lysosomal degradation, is induced to ensure survival under starvation and other stresses. A selective autophagic pathway for 60S ribosomal subunits elicited by nitrogen starvation in yeast—ribophagy—was recently described and requires the Ubp3-Bre5 deubiquitylating enzyme. This discovery implied that an E3 ligases act upstream, whether inhibiting the process or providing an initial required signal. In this paper, we show that Ltn1/Rkr1, a 60S ribosome-associated E3 implicated in translational surveillance, acts as an inhibitor of 60S ribosomal subunit ribophagy and is antagonized by Ubp3. The ribosomal protein Rpl25 is a relevant target. Its ubiquitylation is Ltn1 dependent and Ubp3 reversed, and mutation of its ubiquitylation site rendered ribophagy less dependent on Ubp3. Consistently, the expression of Ltn1—but not Ubp3—rapidly decreased after starvation, presumably to allow ribophagy to proceed. Thus, Ltn1 and Ubp3-Bre5 likely contribute to adapt ribophagy activity to both nutrient supply and protein translation.  相似文献   
5.
The PI 3-kinase (PI 3-K) signaling pathway is essential for Schwann cell myelination. Here we have characterized PI 3-K effectors activated during myelination by probing myelinating cultures and developing nerves with an antibody that recognizes phosphorylated substrates for this pathway. We identified a discrete number of phospho-proteins including the S6 ribosomal protein (S6rp), which is down-regulated at the onset of myelination, and N-myc downstream-regulated gene-1 (NDRG1), which is up-regulated strikingly with myelination. We show that type III Neuregulin1 on the axon is the primary activator of S6rp, an effector of mTORC1. In contrast, laminin-2 in the extracellular matrix (ECM), signaling through the α6β4 integrin and Sgk1 (serum and glucocorticoid-induced kinase 1), drives phosphorylation of NDRG1 in the Cajal bands of the abaxonal compartment. Unexpectedly, mice deficient in α6β4 integrin signaling or Sgk1 exhibit hypermyelination during development. These results identify functionally and spatially distinct PI 3-K pathways: an early, pro-myelinating pathway driven by axonal Neuregulin1 and a later-acting, laminin–integrin-dependent pathway that negatively regulates myelination.  相似文献   
6.
Plasma lipidome is now increasingly recognized as a potentially important marker of chronic diseases, but the exact extent of its contribution to the interindividual phenotypic variability in family studies is unknown. Here, we used the rich data from the ongoing San Antonio Family Heart Study (SAFHS) and developed a novel statistical approach to quantify the independent and additive value of the plasma lipidome in explaining metabolic syndrome (MS) variability in Mexican American families recruited in the SAFHS. Our analytical approach included two preprocessing steps: principal components analysis of the high-resolution plasma lipidomics data and construction of a subject-subject lipidomic similarity matrix. We then used the Sequential Oligogenic Linkage Analysis Routines software to model the complex family relationships, lipidomic similarities, and other important covariates in a variance components framework. Our results suggested that even after accounting for the shared genetic influences, indicators of lipemic status (total serum cholesterol, TGs, and HDL cholesterol), and obesity, the plasma lipidome independently explained 22% of variability in the homeostatic model of assessment-insulin resistance trait and 16% to 22% variability in glucose, insulin, and waist circumference. Our results demonstrate that plasma lipidomic studies can additively contribute to an understanding of the interindividual variability in MS.  相似文献   
7.
Over the past several years, considerable progress has been made in the development of gene therapy as a therapeutic strategy for a variety of inherited metabolic diseases, including neuropathic lysosomal storage disorders (LSDs). The premise of gene therapy for this group of diseases is borne of findings that genetic modification of a subset of cells can provide a more global benefit by virtue of the ability of the secreted lysosomal enzymes to effect cross-correction of adjacent and distal cells. Preclinical studies in small and large animal models of these disorders support the application of either a direct in vivo approach using recombinant adeno-associated viral vectors or an ex vivo strategy using lentiviral vector-modified hematopoietic stem cells to correct the neurological component of these diseases. Early clinical studies utilizing both approaches have begun or are in late-stage planning for a small number of neuropathic LSDs. Although initial indications from these studies are encouraging, it is evident that second-generation vectors that exhibit a greater safety profile and transduction activity may be required before this optimism can be fully realized. Here, I review recent progress and the remaining challenges to treat the neurological aspects of various LSDs using this therapeutic paradigm.  相似文献   
8.
In these studies, the role of ceramide-1-phosphate (C1P) in the wound-healing process was investigated. Specifically, fibroblasts isolated from mice with the known anabolic enzyme for C1P, ceramide kinase (CERK), ablated (CERK−/− mice) and their wild-type littermates (CERK+/+) were subjected to in vitro wound-healing assays. Simulation of mechanical trauma of a wound by scratching a monolayer of fibroblasts from CERK+/+ mice demonstrated steadily increasing levels of arachidonic acid in a time-dependent manner in stark contrast to CERK−/− fibroblasts. This observed difference was reflected in scratch-induced eicosanoid levels. Similar, but somewhat less intense, changes were observed in a more complex system utilizing skin biopsies obtained from CERK-null mice. Importantly, C1P levels increased during the early stages of human wound healing correlating with the transition from the inflammatory stage to the peak of the fibroplasia stage (e.g., proliferation and migration of fibroblasts). Finally, the loss of proper eicosanoid response translated into an abnormal migration pattern for the fibroblasts isolated from CERK−/−. As the proper migration of fibroblasts is one of the necessary steps of wound healing, these studies demonstrate a novel requirement for the CERK-derived C1P in the proper healing response of wounds.  相似文献   
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