全文获取类型
收费全文 | 5116篇 |
免费 | 463篇 |
专业分类
5579篇 |
出版年
2023年 | 10篇 |
2022年 | 57篇 |
2021年 | 75篇 |
2020年 | 40篇 |
2019年 | 72篇 |
2018年 | 77篇 |
2017年 | 74篇 |
2016年 | 140篇 |
2015年 | 233篇 |
2014年 | 282篇 |
2013年 | 332篇 |
2012年 | 395篇 |
2011年 | 355篇 |
2010年 | 249篇 |
2009年 | 248篇 |
2008年 | 325篇 |
2007年 | 311篇 |
2006年 | 328篇 |
2005年 | 329篇 |
2004年 | 303篇 |
2003年 | 237篇 |
2002年 | 245篇 |
2001年 | 65篇 |
2000年 | 44篇 |
1999年 | 55篇 |
1998年 | 83篇 |
1997年 | 42篇 |
1996年 | 39篇 |
1995年 | 37篇 |
1994年 | 45篇 |
1993年 | 44篇 |
1992年 | 36篇 |
1991年 | 31篇 |
1990年 | 51篇 |
1989年 | 30篇 |
1988年 | 28篇 |
1987年 | 17篇 |
1986年 | 18篇 |
1985年 | 26篇 |
1984年 | 21篇 |
1983年 | 15篇 |
1982年 | 14篇 |
1981年 | 15篇 |
1980年 | 13篇 |
1979年 | 10篇 |
1978年 | 12篇 |
1976年 | 14篇 |
1974年 | 6篇 |
1973年 | 7篇 |
1971年 | 7篇 |
排序方式: 共有5579条查询结果,搜索用时 0 毫秒
151.
Eva Veronesi Frank Antony Simon Gubbins Nick Golding Alison Blackwell Peter PC. Mertens Joe Brownlie Karin E. Darpel Philip S. Mellor Simon Carpenter 《PloS one》2013,8(8)
Background
Culicoides biting midges (Diptera: Ceratopogonidae) are the biological vectors of globally significant arboviruses of livestock including bluetongue virus (BTV), African horse sickness virus (AHSV) and the recently emerging Schmallenberg virus (SBV). From 2006–2009 outbreaks of BTV in northern Europe inflicted major disruption and economic losses to farmers and several attempts were made to implicate Palaearctic Culicoides species as vectors. Results from these studies were difficult to interpret as they used semi-quantitative RT-PCR (sqPCR) assays as the major diagnostic tool, a technique that had not been validated for use in this role. In this study we validate the use of these assays by carrying out time-series detection of BTV RNA in two colony species of Culicoides and compare the results with the more traditional isolation of infectious BTV on cell culture.Methodology/Principal Findings
A BTV serotype 1 strain mixed with horse blood was fed to several hundred individuals of Culicoides sonorensis (Wirth & Jones) and C. nubeculosus (Mg.) using a membrane-based assay and replete individuals were then incubated at 25°C. At daily intervals 25 Culicoides of each species were removed from incubation, homogenised and BTV quantified in each individual using sqPCR (Cq values) and virus isolation on a KC-C. sonorensis embryonic cell line, followed by antigen enzyme-linked immunosorbent assay (ELISA). In addition, comparisons were also drawn between the results obtained with whole C. sonorensis and with individually dissected individuals to determine the level of BTV dissemination.Conclusions/Significance
Cq values generated from time-series infection experiments in both C. sonorensis and C. nubeculosus confirmed previous studies that relied upon the isolation and detection of infectious BTV. Implications on the testing of field-collected Culicoides as potential virus vectors by PCR assays and the use of such assays as front-line tools for use in diagnostic laboratories in this role are discussed. 相似文献152.
Abimbola A. Onasoga-Jarvis Karin Leiderman Aaron L. Fogelson Michael Wang Marilyn J. Manco-Johnson Jorge A. Di Paola Keith B. Neeves 《PloS one》2013,8(11)
Clinical evidence suggests that individuals with factor VIII (FVIII) deficiency (hemophilia A) are protected against venous thrombosis, but treatment with recombinant proteins can increase their risk for thrombosis. In this study we examined the dynamics of thrombus formation in individuals with hemophilia A and their response to replacement and bypass therapies under venous flow conditions. Fibrin and platelet accumulation were measured in microfluidic flow assays on a TF-rich surface at a shear rate of 100 s−1. Thrombin generation was calculated with a computational spatial-temporal model of thrombus formation. Mild FVIII deficiencies (5–30% normal levels) could support fibrin fiber formation, while severe (<1%) and moderate (1–5%) deficiencies could not. Based on these experimental observations, computational calculations estimate an average thrombin concentration of ∼10 nM is necessary to support fibrin formation under flow. There was no difference in fibrin formation between severe and moderate deficiencies, but platelet aggregate size was significantly larger for moderate deficiencies. Computational calculations estimate that the local thrombin concentration in moderate deficiencies is high enough to induce platelet activation (>1 nM), but too low to support fibrin formation (<10 nM). In the absence of platelets, fibrin formation was not supported even at normal FVIII levels, suggesting platelet adhesion is necessary for fibrin formation. Individuals treated by replacement therapy, recombinant FVIII, showed normalized fibrin formation. Individuals treated with bypass therapy, recombinant FVIIa, had a reduced lag time in fibrin formation, as well as elevated fibrin accumulation compared to healthy controls. Treatment of rFVIIa, but not rFVIII, resulted in significant changes in fibrin dynamics that could lead to a prothrombotic state. 相似文献
153.
Tatiana Mikhailyuk Andreas Holzinger Petro Tsarenko Karin Glaser Eduard Demchenko Ulf Karsten 《Journal of phycology》2020,56(3):671-686
Several strains of terrestrial algae isolated from biological soil crusts in Germany and Ukraine were identified by morphological methods as the widely distributed species Dictyosphaerium minutum (=Dictyosphaerium chlorelloides). Investigation of the phylogeny showed their position unexpectedly outside of Chlorellaceae (Trebouxiophyceae) and distantly from Chlorella chlorelloides, to which this taxon was attributed after revision of the genus Chlorella based on an integrative approach. SSU rRNA phylogeny determined the position of our strains inside a clade recently described as a new genus of the cryptic alga Xerochlorella olmiae isolated from desert biological soil crusts in the United States. Investigation of the morphology of the authentic strain of X. olmiae showed Dictyosphaerium-like morphology, as well as some other characters, common for our strains and morphospecies D. minutum. The latter alga was described as terrestrial and subsequently united with the earlier described aquatic representative D. chlorelloides because of their similar morphology. The revision of Chlorella mentioned above provided only one aquatic strain (D. chlorelloides), which determined its position in the genus. But terrestrial strains of the morphospecies were not investigated phylogenetically. Our study showed that the terrestrial D. minutum is not related to the morphologically similar D. chlorelloides (=Chlorella chlorelloides, Chlorellaceae), and instead represented a separate lineage in the Trebouxiophyceae, recently described as genus Xerochlorella. Therefore, revision of Xerochlorella is proposed, including nomenclatural combinations, epitypifications, and emendations of two species: X. minuta and X. dichotoma. New characters of the genus based on investigation of morphology and ultrastructure were determined. 相似文献
154.
Annique C. Dombrowsky Karin Burger Ann-Kristin Porth Marlon Stein Martin Dierolf Benedikt Gnther Klaus Achterhold Bernhard Gleich Annette Feuchtinger Stefan Bartzsch Elke Beyreuther Stephanie E. Combs Franz Pfeiffer Jan J. Wilkens Thomas E. Schmid 《Radiation and environmental biophysics》2020,59(1):111-120
Microbeam radiation therapy (MRT), a preclinical form of spatially fractionated radiotherapy, uses an array of microbeams of hard synchrotron X-ray radiation. Recently, compact synchrotron X-ray sources got more attention as they provide essential prerequisites for the translation of MRT into clinics while overcoming the limited access to synchrotron facilities. At the Munich compact light source (MuCLS), one of these novel compact X-ray facilities, a proof of principle experiment was conducted applying MRT to a xenograft tumor mouse model. First, subcutaneous tumors derived from the established squamous carcinoma cell line FaDu were irradiated at a conventional X-ray tube using broadbeam geometry to determine a suitable dose range for the tumor growth delay. For irradiations at the MuCLS, FaDu tumors were irradiated with broadbeam and microbeam irradiation at integral doses of either 3 Gy or 5 Gy and tumor growth delay was measured. Microbeams had a width of 50 µm and a center-to-center distance of 350 µm with peak doses of either 21 Gy or 35 Gy. A dose rate of up to 5 Gy/min was delivered to the tumor. Both doses and modalities delayed the tumor growth compared to a sham-irradiated tumor. The irradiated area and microbeam pattern were verified by staining of the DNA double-strand break marker γH2AX. This study demonstrates for the first time that MRT can be successfully performed in vivo at compact inverse Compton sources. 相似文献
155.
Pedro M. D. Moreno Sylvain Geny Y. Vladimir Pabon Helen Bergquist Eman M. Zaghloul Cristina S. J. Rocha Iulian I. Oprea Burcu Bestas Samir EL Andaloussi Per T. J?rgensen Erik B. Pedersen Karin E. Lundin Rula Zain Jesper Wengel C. I. Edvard Smith 《Nucleic acids research》2013,41(5):3257-3273
In spite of the many developments in synthetic oligonucleotide (ON) chemistry and design, invasion into double-stranded DNA (DSI) under physiological salt and pH conditions remains a challenge. In this work, we provide a new ON tool based on locked nucleic acids (LNAs), designed for strand invasion into duplex DNA (DSI). We thus report on the development of a clamp type of LNA ON—bisLNA—with capacity to bind and invade into supercoiled double-stranded DNA. The bisLNA links a triplex-forming, Hoogsteen-binding, targeting arm with a strand-invading Watson–Crick binding arm. Optimization was carried out by varying the number and location of LNA nucleotides and the length of the triplex-forming versus strand-invading arms. Single-strand regions in target duplex DNA were mapped using chemical probing. By combining design and increase in LNA content, it was possible to achieve a 100-fold increase in potency with 30% DSI at 450 nM using a bisLNA to plasmid ratio of only 21:1. Although this first conceptual report does not address the utility of bisLNA for the targeting of DNA in a chromosomal context, it shows bisLNA as a promising candidate for interfering also with cellular genes. 相似文献
156.
Verónica G. Rodriguez Moncalvo Sabrina S. Burmeister Karin S. Pfennig 《Journal of comparative physiology. A, Neuroethology, sensory, neural, and behavioral physiology》2013,199(8):681-691
Monoamines are important neuromodulators that respond to social cues and that can, in turn, modify social responses. Yet we know very little about the ontogeny of monoaminergic systems and whether they contribute to the development of social behavior. Anurans are an excellent model for studying the development of social behavior because one of its primary components, phonotaxis, is expressed early in life. To examine the effect of social signals on monoamines early in ontogeny, we presented juvenile Mexican spadefoot toads (Spea multiplicata) with a male mating call or no sound and measured norepinephrine, epinephrine, dopamine, serotonin, and a serotonin metabolite, across the brain using high-pressure liquid chromatography. Our results demonstrate that adult-like monoaminergic systems are in place shortly after metamorphosis. Perhaps more interestingly, we found that mating calls increased the level of monoamines in the juvenile tegmentum, a midbrain region involved in sensory-motor integration and that contributes to brain arousal and attention. We saw no such increase in the auditory midbrain or in forebrain regions. We suggest that changes in monoamine levels in the juvenile tegmentum may reflect the effects of social signals on arousal state and could contribute to context-dependent modulation of social behavior. 相似文献
157.
Karin Rybka Siobhan E. Toal Daniel J. Verbaro Daniel Mathieu Harald Schwalbe Reinhard Schweitzer‐Stenner 《Proteins》2013,81(6):968-983
In the preceding paper, we found that ensembles of tripeptides with long or bulky chains can include up to 20% of various turns. Here, we determine the structural and thermodynamic characteristics of GxG peptides with short polar and/or ionizable central residues (D, N, C), whose conformational distributions exhibit higher than average percentage (>20%) of turn conformations. To probe the side‐chain conformations of these peptides, we determined the 3J(Hα,Hβ) coupling constants and derived the population of three rotamers with χ1‐angles of ?60°, 180° and 60°, which were correlated with residue propensities by DFT‐calculations. For protonated GDG, the rotamer distribution provides additional evidence for asx‐turns. A comparison of vibrational spectra and NMR coupling constants of protonated GDG, ionized GDG, and the protonated aspartic acid dipeptide revealed that side chain protonation increases the pPII content at the expense of turn populations. The charged terminal groups, however, have negligible influence on the conformational properties of the central residue. Like protonated GDG, cationic GCG samples asx‐turns to a significant extent. The temperature dependence of the UVCD spectra and 3J(HNHα) constants suggest that the turn populations of GDG and GNG are practically temperature‐independent, indicating enthalpic and entropic stabilization. The temperature‐independent J‐coupling and UVCD spectra of GNG require a three‐state model. Our results indicate that short side chains with hydrogen bonding capability in GxG segments of proteins may serve as hinge regions for establishing compact structures of unfolded proteins and peptides. Proteins 2013. © 2012 Wiley Periodicals, Inc. 相似文献
158.
Wibke Singer Annalisa Zuccotti Mirko Jaumann Sze Chim Lee Rama Panford-Walsh Hao Xiong Ulrike Zimmermann Christoph Franz Hyun-Soon Geisler Iris Köpschall Karin Rohbock Ksenya Varakina Sandrine Verpoorten Thomas Reinbothe Thomas Schimmang Lukas Rüttiger Marlies Knipper 《Molecular neurobiology》2013,47(1):261-279
Increasing evidence shows that hearing loss is a risk factor for tinnitus and hyperacusis. Although both often coincide, a causal relationship between tinnitus and hyperacusis has not been shown. Currently, tinnitus and hyperacusis are assumed to be caused by elevated responsiveness in subcortical circuits. We examined both the impact of different degrees of cochlear damage and the influence of stress priming on tinnitus induction. We used (1) a behavioral animal model for tinnitus designed to minimize stress, (2) ribbon synapses in inner hair cells (IHCs) as a measure for deafferentation, (3) the integrity of auditory brainstem responses (ABR) to detect differences in stimulus-evoked neuronal activity, (4) the expression of the activity-regulated cytoskeletal protein, Arc, to identify long-lasting changes in network activity within the basolateral amygdala (BLA), hippocampal CA1, and auditory cortex (AC), and (5) stress priming to investigate the influence of corticosteroid on trauma-induced brain responses. We observed that IHC ribbon loss (deafferentation) leads to tinnitus when ABR functions remain reduced and Arc is not mobilized in the hippocampal CA1 and AC. If, however, ABR waves are functionally restored and Arc is mobilized, tinnitus does not occur. Both central response patterns were found to be independent of a profound threshold loss and could be shifted by the corticosterone level at the time of trauma. We, therefore, discuss the findings in the context of a history of stress that can trigger either an adaptive or nonadaptive brain response following injury. 相似文献
159.
Karin E. van Straaten Jong Bum Ko Rajendra Jagdhane Shazia Anjum David R. J. Palmer David A. R. Sanders 《The Journal of biological chemistry》2013,288(47):34121-34130
NtdA from Bacillus subtilis is a sugar aminotransferase that catalyzes the pyridoxal phosphate-dependent equatorial transamination of 3-oxo-α-d-glucose 6-phosphate to form α-d-kanosamine 6-phosphate. The crystal structure of NtdA shows that NtdA shares the common aspartate aminotransferase fold (Type 1) with residues from both monomers forming the active site. The crystal structures of NtdA alone, co-crystallized with the product α-d-kanosamine 6-phosphate, and incubated with the amine donor glutamate reveal three key structures in the mechanistic pathway of NtdA. The structure of NtdA alone reveals the internal aldimine form of NtdA with the cofactor pyridoxal phosphate covalently attached to Lys-247. The addition of glutamate results in formation of pyridoxamine phosphate. Co-crystallization with kanosamine 6-phosphate results in the formation of the external aldimine. Only α-d-kanosamine 6-phosphate is observed in the active site of NtdA, not the β-anomer. A comparison of the structure and sequence of NtdA with other sugar aminotransferases enables us to propose that the VIβ family of aminotransferases should be divided into subfamilies based on the catalytic lysine motif. 相似文献
160.
Viviana Ulloa María García‐Robles Fernando Martínez Katterine Salazar Karin Reinicke Fernando Pérez David F. Godoy Alejandro S. Godoy Francisco Nualart 《Journal of neurochemistry》2013,127(3):403-414
In vitro and in vivo studies suggest that the basolateral membrane of choroid plexus cells, which is in contact with blood vessels, is involved in the uptake of the reduced form of vitamin C, ascorbic acid (AA), through the sodium‐vitamin C cotransporter, (SVCT2). Moreover, very low levels of vitamin C were observed in the brains of SVCT2‐null mice. The oxidized form of vitamin C, dehydroascorbic acid (DHA), is incorporated through the facilitative glucose transporters (GLUTs). In this study, the contribution of SVCT2 and GLUT1 to vitamin C uptake in human choroid plexus papilloma (HCPP) cells in culture was examined. Both the functional activity and the kinetic parameters of GLUT1 and SVCT2 in cells isolated from HCPP were observed. Finally, DHA uptake by GLUT1 in choroid plexus cells was assessed in the presence of phorbol‐12‐myristate‐13‐acetate (PMA)‐activated human neutrophils. A marked increase in vitamin C uptake by choroid plexus cells was observed that was associated with superoxide generation and vitamin C oxidation (bystander effect). Thus, vitamin C can be incorporated by epithelial choroid plexus papilloma cells using the basolateral polarization of SVCT2 and GLUT1. This mechanism may be amplified with neutrophil infiltration (inflammation) of choroid plexus tumors.