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991.
Schock DM Ruthig GR Collins JP Kutz SJ Carrière S Gau RJ Veitch AM Larter NC Tate DP Guthrie G Allaire DG Popko RA 《Diseases of aquatic organisms》2010,92(2-3):231-240
Pathogens can cause serious declines in host species, and knowing where pathogens associated with host declines occur facilitates understanding host-pathogen ecology. Suspected drivers of global amphibian declines include infectious diseases, with 2 pathogens in particular, Batrachochytrium dendrobatidis (Bd) and ranaviruses, causing concern. We explored the host range and geographic distribution of Bd and ranaviruses in the Taiga Plains ecoregion of the Northwest Territories, Canada, in 2007 and 2008. Both pathogens were detected, greatly extending their known geographic distributions. Ranaviruses were widespread geographically, but found only in wood frogs. In contrast, Bd was found at a single site, but was detected in all 3 species of amphibians in the survey area (wood frogs, boreal chorus frogs, western toads). The presence of Bd in the Northwest Territories is not congruent with predicted distributions based on niche models, even though findings from other studies at northern latitudes are consistent with those same models. Unexpectedly, we also found evidence that swabs routinely used to collect samples for Bd screening detected fewer infections than toe clips. Our use and handling of the swabs was consistent with other studies, and the cause of the apparent lack of integrity of swabs is unknown. The ranaviruses detected in our study were confirmed to be Frog Virus 3 by sequence analysis of a diagnostic 500 bp region of the major capsid protein gene. It is unknown whether Bd or ranaviruses are recent arrivals to the Canadian north. However, the genetic analyses required to answer that question can inform larger debates about the origin of Bd in North America as well as the potential effects of climate change and industrial development on the distributions of these important amphibian pathogens. 相似文献
992.
Geoffrey E. Woodard Ning-Na Huang Hyeseon Cho Toru Miki Gregory G. Tall John H. Kehrl 《Molecular and cellular biology》2010,30(14):3519-3530
In model organisms, resistance to inhibitors of cholinesterase 8 (Ric-8), a G protein α (Gα) subunit guanine nucleotide exchange factor (GEF), functions to orient mitotic spindles during asymmetric cell divisions; however, whether Ric-8A has any role in mammalian cell division is unknown. We show here that Ric-8A and Gαi function to orient the metaphase mitotic spindle of mammalian adherent cells. During mitosis, Ric-8A localized at the cell cortex, spindle poles, centromeres, central spindle, and midbody. Pertussis toxin proved to be a useful tool in these studies since it blocked the binding of Ric-8A to Gαi, thus preventing its GEF activity for Gαi. Linking Ric-8A signaling to mammalian cell division, treatment of cells with pertussis toxin, reduction of Ric-8A expression, or decreased Gαi expression similarly affected metaphase cells. Each treatment impaired the localization of LGN (GSPM2), NuMA (microtubule binding nuclear mitotic apparatus protein), and dynein at the metaphase cell cortex and disturbed integrin-dependent mitotic spindle orientation. Live cell imaging of HeLa cells expressing green fluorescent protein-tubulin also revealed that reduced Ric-8A expression prolonged mitosis, caused occasional mitotic arrest, and decreased mitotic spindle movements. These data indicate that Ric-8A signaling leads to assembly of a cortical signaling complex that functions to orient the mitotic spindle.The cortical capture of astral microtubules is essential to generate the forces needed for mitotic spindle positioning for both symmetric and asymmetric cell divisions (23, 29). Failure to either capture astral microtubules or the inappropriate application of pulling forces adversely affects mitotic spindle orientation, and can impede embryogenesis and alter cell fate decisions. Studies examining mitotic spindle orientation in Drosophila embryonic and larval neuroblasts have identified two critical pathways, the Gα/Pins/Mud pathway and the Pins/Dlg/Khc73 pathway (29). The heterotrimeric G-protein α subunit (Gα), Pins (Partner-of-Inscuteable), and Mud (Mushroom body defect) are members of an evolutionarily conserved noncanonical G-protein signaling pathway, which form a tripartite protein complex linked to the apical Par complex by the adapter protein Inscuteable (29, 37). Reducing the level of Gαi, Pins, or Mud prevents neuroblast mitotic spindle alignment. A second spindle orientation pathway involves Pins, the tumor suppressor Discs large (Dlg) and the microtubule plus-end-directed kinesin heavy chain 73 (Khc73). Khc73 binds Dlg and coimmunoprecipitates with Pins. Khc73 localized to astral microtubules can induce Pins-Dlg cortical polarity (27).In canonical G-protein signaling pathways, the binding of ligand to a seven-transmembrane receptor triggers a heterotrimeric G-protein α subunit (Gα) to exchange GTP for GDP, resulting in the dissociation of the Gα subunit from its associated Gβγ heterodimer (12, 20). This exposes interactive sites in the Gα and Gβγ subunits, allowing their binding to and activation of downstream effectors. Since Gα subunits possess an intrinsic GTPase activity, GTP hydrolysis leads to the reassembly of heterotrimeric G protein causing signaling to cease. In noncanonical G-protein signaling the seven-transmembrane receptor is replaced by an intracellular guanine nucleotide exchange factor, such as Ric-8 (37). In studies in Drosophila and Caenorhabditis elegans Ric-8 has been shown to positively regulate Gαi activity and is essential for asymmetric cell divisions (1, 2, 5, 8, 11, 36). Although initially characterized as a guanine nucleotide exchange factor (GEF) for isolated Gαsubunits, more recent biochemical studies have shown that Ric-8A (the mammalian equivalent of Ric-8) also acts on a complex of GDP-Gαi, the mammalian Pins homolog LGN, and NuMA (nuclear mitotic apparatus protein; the mammalian equivalent of Mud) catalytically releasing GTP-Gαi and causing liberation of NuMA from LGN (30, 31). Ric-8A can also catalyze guanine nucleotide exchange on Gαi1 bound to the GPR/GoLoco exchange inhibitor AGS3, a paralog of LGN (33). During mitosis the N-terminal portion of LGN binds NuMA and the C-terminal domain binds GDP-Gαi and the trimolecular complex localizes to the cell cortex, where the dynamic release of NuMA from LGN may regulate aster microtubule pulling during cell division (3, 9, 10, 22).In the present study we examined the role of Ric-8A in mitotic spindle orientation in adherent cells and in polarized MDCK cells. In nonpolarized adherent cells cell such as HeLa, integrin mediated cell-substrate adhesion orients the mitotic spindle parallel to the substratum, and thereby both daughter cells remain attached. This requires the actin cytoskeleton, astral microtubules, the microtubule plus end tracking protein EB1, myosin X, cdc42, LIM kinase 1, and phosphatidylinositol(3,4,5)-triphosphate (PIP3) (13, 18, 32, 34, 35). PIP3 may direct dynein/dynactin-dependent pulling forces on the spindle midcortex to orient the mitotic spindle (34). In polarized cells such as Madin-Darby canine kidney (MDCK) cells, the mitotic spindle is constrained by the topology of the cell and cortical cues provided by adherens junctions (24). In contrast to HeLa cells these cues are insensitive to phosphatidylinositol 3-kinase (PI3K) inhibition, which blocks the generation of PIP3 (34). We found that inhibiting either Ric-8A or Gαi expression impairs the orientation of the metaphase mitotic spindle in HeLa cells and pertussis toxin, which blocks Ric-8A triggered nucleotide exchange, disrupts the normal mitotic spindle alignment of both HeLa and MDCK cells. Impairment of Ric-8A expression or function inhibits the localization of Gαi1, LGN, NuMA, and dynein to the metaphase cortex opposite the spindle poles. 相似文献
993.
An individual-based model (IbM) for bacterial adaptation and evolution, COSMIC-Rules, has been employed to simulate interactions of virtual temperate bacteriophages (phages) and their bacterial hosts. Outcomes of infection mimic those of a phage such as lambda, which can enter either the lytic or lysogenic cycle, depending on the nutritional status of the host. Infection of different hosts possessing differing restriction and modification systems is also simulated. Phages restricted upon infection of one restricting host can be adapted (by host-controlled modification of the phage genome) and subsequently propagate with full efficiency on this host. However, such ability is lost if the progeny phages are passaged through a new host with a different restriction and modification system before attempted re-infection of the original restrictive host. The simulations show that adaptation and re-adaptation to a particular host-controlled restriction and modification system result in lower efficiency and delayed lysis of bacterial cells compared with infection of non-restricting host bacteria. 相似文献
994.
Internalization of Flax Rust Avirulence Proteins into Flax and Tobacco Cells Can Occur in the Absence of the Pathogen 总被引:1,自引:0,他引:1
Maryam Rafiqi Pamela H.P. Gan Michael Ravensdale Gregory J. Lawrence Jeffrey G. Ellis David A. Jones Adrienne R. Hardham Peter N. Dodds 《The Plant cell》2010,22(6):2017-2032
Translocation of pathogen effector proteins into the host cell cytoplasm is a key determinant for the pathogenicity of many bacterial and oomycete plant pathogens. A number of secreted fungal avirulence (Avr) proteins are also inferred to be delivered into host cells, based on their intracellular recognition by host resistance proteins, including those of flax rust (Melampsora lini). Here, we show by immunolocalization that the flax rust AvrM protein is secreted from haustoria during infection and accumulates in the haustorial wall. Five days after inoculation, the AvrM protein was also detected within the cytoplasm of a proportion of plant cells containing haustoria, confirming its delivery into host cells during infection. Transient expression of secreted AvrL567 and AvrM proteins fused to cerulean fluorescent protein in tobacco (Nicotiana tabacum) and flax cells resulted in intracellular accumulation of the fusion proteins. The rust Avr protein signal peptides were functional in plants and efficiently directed fused cerulean into the secretory pathway. Thus, these secreted effectors are internalized into the plant cell cytosol in the absence of the pathogen, suggesting that they do not require a pathogen-encoded transport mechanism. Uptake of these proteins is dependent on signals in their N-terminal regions, but the primary sequence features of these uptake regions are not conserved between different rust effectors. 相似文献
995.
Yan F Wu X Crawford M Duan W Wilding EE Gao L Nana-Sinkam SP Villalona-Calero MA Baiocchi RA Otterson GA 《Methods (San Diego, Calif.)》2010,52(4):281-286
Clinical trials and correlative laboratory research are increasingly reliant upon archived paraffin-embedded samples. Therefore, the proper processing of biological samples is an important step to sample preservation and for downstream analyses like the detection of a wide variety of targets including micro RNA, DNA and proteins. This paper analyzed the question whether routine fixation of cells and tissues in 10% buffered formalin is optimal for in situ and solution phase analyses by comparing this fixative to a variety of cross linking and alcohol (denaturing) fixatives. We examined the ability of nine commonly used fixative regimens to preserve cell morphology and DNA/RNA/protein quality for these applications. Epstein-Barr virus (EBV) and bovine papillomavirus (BPV)-infected tissues and cells were used as our model systems. Our evaluation showed that the optimal fixative in cell preparations for molecular hybridization techniques was "gentle" fixative with a cross-linker such as paraformaldehyde or a short incubation in 10% buffered formalin. The optimal fixatives for tissue were either paraformaldehyde or low concentration of formalin (5% of formalin). Methanol was the best of the non cross-linking fixatives for in situ hybridization and immunohistochemistry. For PCR-based detection of DNA or RNA, some denaturing fixatives like acetone and methanol as well as "gentle" cross-linking fixatives like paraformaldehyde out-performed other fixatives. Long term fixation was not proposed for DNA/RNA-based assays. The typical long-term fixation of cells and tissues in 10% buffered formalin is not optimal for combined analyses by in situ hybridization, immunohistochemistry, or--if one does not have unfixed tissues--solution phase PCR. Rather, we recommend short term less intense cross linking fixation if one wishes to use the same cells/tissue for in situ hybridization, immunohistochemistry, and solution phase PCR. 相似文献
996.
Stefan Schmid Marilyn Moffat Gregory M. Gutierrez 《Journal of electromyography and kinesiology》2010,20(6):1075-1081
During sporting events, injured athletes often return to competition after icing because of the reduction in pain. Although some controversy exists, several studies suggest that cryotherapy causes a decrease in muscle activity, which may lead to a higher risk of injury upon return to play. The purpose of this study was to investigate the effect of a 20-min knee joint cryotherapy application on the electromyographic activity of leg muscles during a single-leg drop jump in twenty healthy subjects, randomly assigned to an experimental and a control group. After the pre-tests, a crushed-ice bag was applied to the knee joint of the experimental group subjects for 20 min, while the control group subjects rested for 20 min. All subjects were retested immediately after this period and retested again after another 20 min of rest. Average electromyographic activity and ground contact time were calculated for the pre- and post-test sessions. Decreases in electromyographic activity of the lower extremity musculature were found in pre-activation, eccentric (braking), and concentric (push-off) phases immediately after the icing, and after 20 min of rest. The results lend support to the suggestion that cryotherapy during sporting events may place the individuals in a vulnerable position. 相似文献
997.
Rita Barallon Steven R. Bauer John Butler Amanda Capes-Davis Wilhelm G. Dirks Eugene Elmore Manohar Furtado Margaret C. Kline Arihiro Kohara Georgyi V. Los Roderick A. F. MacLeod John R. W. Masters Mark Nardone Roland M. Nardone Raymond W. Nims Paul J. Price Yvonne A. Reid Jaiprakash Shewale Gregory Sykes Anton F. Steuer Douglas R. Storts Jim Thomson Zenobia Taraporewala Christine Alston-Roberts Liz Kerrigan 《In vitro cellular & developmental biology. Animal》2010,46(9):727-732
Cell misidentification and cross-contamination have plagued biomedical research for as long as cells have been employed as research tools. Examples of misidentified cell lines continue to surface to this day. Efforts to eradicate the problem by raising awareness of the issue and by asking scientists voluntarily to take appropriate actions have not been successful. Unambiguous cell authentication is an essential step in the scientific process and should be an inherent consideration during peer review of papers submitted for publication or during review of grants submitted for funding. In order to facilitate proper identity testing, accurate, reliable, inexpensive, and standardized methods for authentication of cells and cell lines must be made available. To this end, an international team of scientists is, at this time, preparing a consensus standard on the authentication of human cells using short tandem repeat (STR) profiling. This standard, which will be submitted for review and approval as an American National Standard by the American National Standards Institute, will provide investigators guidance on the use of STR profiling for authenticating human cell lines. Such guidance will include methodological detail on the preparation of the DNA sample, the appropriate numbers and types of loci to be evaluated, and the interpretation and quality control of the results. Associated with the standard itself will be the establishment and maintenance of a public STR profile database under the auspices of the National Center for Biotechnology Information. The consensus standard is anticipated to be adopted by granting agencies and scientific journals as appropriate methodology for authenticating human cell lines, stem cells, and tissues. 相似文献
998.
Brett M. Macey Matthew J. Jenny Heidi R. Williams Lindy K. Thibodeaux Marion Beal Jonas S. Almeida Charles Cunningham Annalaura Mancia Gregory W. Warr Erin J. Burge A. Fred Holland Paul S. Gross Sonomi Hikima Karen G. Burnett Louis Burnett Robert W. Chapman 《Comparative biochemistry and physiology. Part A, Molecular & integrative physiology》2010,155(3):341-349
Heavy metals, such as copper, zinc and cadmium, represent some of the most common and serious pollutants in coastal estuaries. In the present study, we used a combination of linear and artificial neural network (ANN) modelling to detect and explore interactions among low-dose mixtures of these heavy metals and their impacts on fundamental physiological processes in tissues of the Eastern oyster, Crassostrea virginica. Animals were exposed to Cd (0.001–0.400 μM), Zn (0.001–3.059 μM) or Cu (0.002–0.787 μM), either alone or in combination for 1 to 27 days. We measured indicators of acid–base balance (hemolymph pH and total CO2), gas exchange (Po2), immunocompetence (total hemocyte counts, numbers of invasive bacteria), antioxidant status (glutathione, GSH), oxidative damage (lipid peroxidation; LPx), and metal accumulation in the gill and the hepatopancreas. Linear analysis showed that oxidative membrane damage from tissue accumulation of environmental metals was correlated with impaired acid–base balance in oysters. ANN analysis revealed interactions of metals with hemolymph acid–base chemistry in predicting oxidative damage that were not evident from linear analyses. These results highlight the usefulness of machine learning approaches, such as ANNs, for improving our ability to recognize and understand the effects of sub-acute exposure to contaminant mixtures. 相似文献
999.
Haffner CD Thomson SA Guo Y Petrov K Larkin A Banker P Schaaf G Dickerson S Gobel J Gillie D Condreay JP Poole C Carpenter T Ulrich J 《Bioorganic & medicinal chemistry letters》2010,20(23):6989-6992
We report the synthesis and in vitro activity of a series of novel substituted N-{3-[(1,1-dioxido-1,2-benzothiazol-3-yl)(phenyl)amino]propyl}benzamide analogs. These analogs showed potent inhibitory activity against Kv1.3. Several demonstrated similar potency to the known Kv1.3 inhibitor PAP-1 when tested under the IonWorks patch clamp assay conditions. Two compounds 13i and 13rr were advanced further as potential tool compounds for in vivo validation studies. 相似文献
1000.
Frédéric J. Zécri Rainer Albert Gregory Landrum Klaus Hinterding Nigel G. Cooke Danilo Guerini Markus Streiff Christian Bruns Barbara Nuesslein-Hildesheim 《Bioorganic & medicinal chemistry letters》2010,20(1):35-37
High throughput screening and hit to lead optimization led to the identification of ‘carene’ as a promising scaffold showing selective S1P1 receptor agonism. In parallel to this work we have established a pharmacophore model for the S1P1 receptor highlighting the minimal structural requirement necessary for potent receptor agonism. 相似文献