全文获取类型
收费全文 | 6630篇 |
免费 | 503篇 |
国内免费 | 2篇 |
专业分类
7135篇 |
出版年
2022年 | 49篇 |
2021年 | 106篇 |
2020年 | 63篇 |
2019年 | 68篇 |
2018年 | 75篇 |
2017年 | 86篇 |
2016年 | 145篇 |
2015年 | 249篇 |
2014年 | 274篇 |
2013年 | 338篇 |
2012年 | 453篇 |
2011年 | 444篇 |
2010年 | 305篇 |
2009年 | 262篇 |
2008年 | 395篇 |
2007年 | 363篇 |
2006年 | 331篇 |
2005年 | 368篇 |
2004年 | 315篇 |
2003年 | 350篇 |
2002年 | 312篇 |
2001年 | 82篇 |
2000年 | 59篇 |
1999年 | 89篇 |
1998年 | 98篇 |
1997年 | 80篇 |
1996年 | 53篇 |
1995年 | 53篇 |
1994年 | 56篇 |
1993年 | 60篇 |
1992年 | 80篇 |
1991年 | 52篇 |
1990年 | 50篇 |
1989年 | 37篇 |
1988年 | 57篇 |
1987年 | 44篇 |
1986年 | 31篇 |
1985年 | 51篇 |
1984年 | 55篇 |
1983年 | 34篇 |
1982年 | 42篇 |
1981年 | 47篇 |
1980年 | 40篇 |
1979年 | 33篇 |
1978年 | 38篇 |
1977年 | 28篇 |
1976年 | 25篇 |
1974年 | 24篇 |
1973年 | 17篇 |
1956年 | 15篇 |
排序方式: 共有7135条查询结果,搜索用时 0 毫秒
991.
992.
Sofie Celen Jan Cleynhens Christophe Deroose Tjibbe de Groot Abdelilah Ibrahimi Rik Gijsbers Zeger Debyser Luc Mortelmans Alfons Verbruggen Guy Bormans 《Bioorganic & medicinal chemistry》2009,17(14):5117-5125
In our aim to develop LacZ reporter probes with a good retention in LacZ expressing cells, we report the synthesis and preliminary evaluation of two carbon-11 labeled β-galactosyl triazoles 1-(β-d-galactopyranosyl)-4-(p-[11C]methoxyphenyl)-1,2,3-triazole ([11C]-6) and 1-(β-d-galactopyranosyl)-4-(6-[11C]methoxynaphthyl)-1,2,3-triazole ([11C]-13). The precursors for the radiolabeling and the non-radioactive analogues (6 and 13) were synthesized using straightforward ‘click’ chemistry. In vitro incubation experiments of 6 with β-galactosidase in the presence of o-nitrophenyl β-d-galactopyranoside (ONPG) showed that the triazolic compound was an inhibitor of β-galactosidase activity. Radiolabeling of both precursors was performed using [11C]methyl iodide as alkylating agent at 70 °C in DMF in the presence of a small amount of base. The log P values were ?0.1 and 1.4, respectively, for [11C]-6 and [11C]-13, the latter therefore being a good candidate for increased cellular uptake via passive diffusion. Biodistribution studies in normal mice showed a good clearance from blood for both tracers. [11C]-6 was mainly cleared via the renal pathway, while the more lipophilic [11C]-13 was excreted almost exclusively via the hepatobiliary system. Despite the lipophilicity of [11C]-13, no brain uptake was observed. Reversed phase HPLC analysis of murine plasma and urine revealed high in vivo stability for both tracers. In vitro evaluation in HEK-293T cells showed an increased cell uptake for the more lipophilic [11C]-13, however, there was no statistically higher uptake in LacZ expressing cells compared to control cells. 相似文献
993.
994.
995.
Ralph Mac Nally rew F. Bennett James R. Thomson James Q. Radford † Guy Unmack Gregory Horrocks Peter A. Vesk 《Diversity & distributions》2009,15(4):720-730
Aim We characterized changes in reporting rates and abundances of bird species over a period of severe rainfall deficiency and increasing average temperatures. We also measured flowering in eucalypts, which support large numbers of nectarivores characteristic of the region.
Location A 30,000-km2 region of northern Victoria, Australia, consisting of limited amounts of remnant native woodlands embedded in largely agricultural landscapes.
Methods There were three sets of monitoring studies, pitched at regional (survey programmes in 1995–97, 2004–05 and 2006–08), landscape (2002–03 and 2006–07) and site (1997–2008 continuously) scales. Bird survey techniques used a standard 2-ha, 20-min count method. We used Bayesian analyses of reporting rates to document statistically changes in the avifauna through time at each spatial scale.
Results Bird populations in the largest remnants of native vegetation (up to 40,000 ha), some of which have been declared as national parks in the past decade, experienced similar declines to those in heavily cleared landscapes. All categories of birds (guilds based on foraging substrate, diet, nest site; relative mobility; geographical distributions) were affected similarly. We detected virtually no bird breeding in the latest survey periods. Eucalypt flowering has declined significantly over the past 12 years of drought.
Main conclusions Declines in the largest woodland remnants commensurate with those in cleared landscapes suggest that reserve systems may not be relied upon to sustain species under climate change. We attribute population declines to low breeding success due to reduced food. Resilience of bird populations in this woodland system might be increased by active management to enhance habitat quality in existing vegetation and restoration of woodland in the more fertile parts of landscapes. 相似文献
Location A 30,000-km
Methods There were three sets of monitoring studies, pitched at regional (survey programmes in 1995–97, 2004–05 and 2006–08), landscape (2002–03 and 2006–07) and site (1997–2008 continuously) scales. Bird survey techniques used a standard 2-ha, 20-min count method. We used Bayesian analyses of reporting rates to document statistically changes in the avifauna through time at each spatial scale.
Results Bird populations in the largest remnants of native vegetation (up to 40,000 ha), some of which have been declared as national parks in the past decade, experienced similar declines to those in heavily cleared landscapes. All categories of birds (guilds based on foraging substrate, diet, nest site; relative mobility; geographical distributions) were affected similarly. We detected virtually no bird breeding in the latest survey periods. Eucalypt flowering has declined significantly over the past 12 years of drought.
Main conclusions Declines in the largest woodland remnants commensurate with those in cleared landscapes suggest that reserve systems may not be relied upon to sustain species under climate change. We attribute population declines to low breeding success due to reduced food. Resilience of bird populations in this woodland system might be increased by active management to enhance habitat quality in existing vegetation and restoration of woodland in the more fertile parts of landscapes. 相似文献
996.
Virginie Le Rolle Guy Carrault Pierre-Yves Richard Philippe Pibarot Louis-Gilles Durand Alfredo I. Hernández 《Acta biotheoretica》2009,57(4):457-478
The ventricular pressure profile is characteristic of the cardiac contraction progress and is useful to evaluate the cardiac
performance. In this contribution, a tissue-level electromechanical model of the left ventricle is proposed, to assist the
interpretation of left ventricular pressure waveforms. The left ventricle has been modeled as an ellipsoid composed of twelve
mechano-hydraulic sub-systems. The asynchronous contraction of these twelve myocardial segments has been represented in order
to reproduce a realistic pressure profiles. To take into account the different energy domains involved, the tissue-level scale
and to facilitate the building of a modular model, multiple formalisms have been used: Bond Graph formalism for the mechano-hydraulic
aspects and cellular automata for the electrical activation. An experimental protocol has been defined to acquire ventricular
pressure signals from three pigs, with different afterload conditions. Evolutionary Algorithms have been used to identify
the model parameters in order to minimize the error between experimental and simulated ventricular pressure signals. Simulation
results show that the model is able to reproduce experimental ventricular pressure. In addition, electro-mechanical activation
times have been determined in the identification process. For example, the maximum electrical activation time is reached,
respectively, 96.5, 139.3 and 131.5 ms for the first, second, and third pigs. These preliminary results are encouraging for
the application of the model on non-invasive data like ECG, arterial pressure or myocardial strain. 相似文献
997.
Mitochondrial Ca2+ transport was initially considered important only in buffering of cytosolic Ca2+ by acting as a “sink” under conditions of Ca2+ overload. The main regulator of ATP production was considered to be the relative concentrations of high energy phosphates. However, work by Denton and McCormack in the 1970s and 1980s showed that free intramitochondrial Ca2+ ([Ca2+]m) activated dehydrogenase enzymes in mitochondria, leading to increased NADH and hence ATP production. This leads them to propose a scheme, subsequently termed a “parallel activation model” whereby increases in energy demand, such as hormonal stimulation or increased workload in muscle, produced an increase in cytosolic [Ca2+] that was relayed by the mitochondrial Ca2+ transporters into the matrix to give an increase in [Ca2+]m. This then stimulated energy production to meet the increased energy demand. With the development of methods for measuring [Ca2+]m in living cells that proved [Ca2+]m changed over a dynamic physiological range rather than simply soaking up excess cytosolic [Ca2+], this model has now gained widespread acceptance. However, work by ourselves and others using targeted probes to measure changes in both [Ca2+] and [ATP] in different cell compartments has revealed variations in the interrelationships between these two in different tissues, suggesting that metabolic regulation by Ca2+ is finely tuned to the demands and function of the individual organ. 相似文献
998.
Subhashini Srinivasan Fozia Mir Jin-Sheng Huang Fadi T. Khasawneh Stephen C.-T. Lam Guy C. Le Breton 《The Journal of biological chemistry》2009,284(24):16108-16117
ADP plays an integral role in the process of hemostasis by signaling through two platelet G-protein-coupled receptors, P2Y1 and P2Y12. The recent use of antagonists against these two receptors has contributed a substantial body of data characterizing the ADP signaling pathways in human platelets. Specifically, the results have indicated that although P2Y1 receptors are involved in the initiation of platelet aggregation, P2Y12 receptor activation appears to account for the bulk of the ADP-mediated effects. Based on this consideration, emphasis has been placed on the development of a new class of P2Y12 antagonists (separate from clopidogrel and ticlopidine) as an approach to the treatment of thromboembolic disorders. The present work examined the molecular mechanisms by which two of these widely used adenosine-based P2Y12 antagonists (2-methylthioadenosine 5′-monophosphate triethylammonium salt (2MeSAMP) and ARC69931MX), inhibit human platelet activation. It was found that both of these compounds raise platelet cAMP to levels that substantially inhibit platelet aggregation. Furthermore, the results demonstrated that this elevation of cAMP did not require Gi signaling or functional P2Y12 receptors but was mediated through activation of a separate G protein-coupled pathway, presumably involving Gs. However, additional experiments revealed that neither 2MeSAMP nor ARC69931MX (cangrelor) increased cAMP through activation of A2a, IP, DP, or EP2 receptors, which are known to couple to Gs. Collectively, these findings indicate that 2MeSAMP and ARC69931MX interact with an unidentified platelet G protein-coupled receptor that stimulates cAMP-mediated inhibition of platelet function. This inhibition is in addition to that derived from antagonism of P2Y12 receptors.Upon damage to the endothelial layer of the blood vessel wall, the underlying subendothelium is exposed to platelets in the blood, initiating a cascade of signaling events resulting in the transformation of “resting” platelets into “activated” platelets (1). One significant characteristic associated with these signaling events is the secretion of ADP from the platelet-dense granules (2). This released ADP acts to further amplify the platelet activation response by interacting with its G-protein-coupled receptors on the platelet surface, namely P2Y1 (coupled to Gq) and P2Y12 (coupled to Gi) (3–5). The consequence of platelet activation through ADP is a conformational change in the platelet membrane glycoprotein αIIbβ3 (6, 7), which then binds to fibrinogen present in the plasma. The binding of fibrinogen with αIIbβ3 on the surface of adjacent platelets results in fibrinogen-platelet cross-linking and the formation of a hemostatic plug at the site of vascular injury (8).Consequently, ADP is thought to play an integral role in the normal process of hemostasis. Of the two ADP-receptor signaling pathways in platelets, evidence has indicated that ADP-mediated P2Y12 signaling appears to play a more prominent role in platelet activation than ADP-mediated P2Y1 signaling (9, 10). For the most part, support for this notion derives from the use of the adenosine-based P2Y12 antagonists (i.e. 2MeSAMP4 and ARC69931MX), which have a much broader inhibitory profile than P2Y1 antagonists (e.g. A3P5P (adenosine-3′-phosphate-5′-phosphate) or MRS2179) (9). Thus, 2MeSAMP and ARC69931MX inhibit platelet aggregation in response to multiple agonists, such as thromboxane A2, collagen, thrombin, etc. (11–13), whereas P2Y1 antagonists do not. On the other hand, this general requirement for P2Y12 signaling seems to be inconsistent with earlier reports indicating that activation of certain platelet receptors (e.g. thromboxane A2 receptor) can cause aggregation through ADP-independent mechanisms (14, 15). Based on this apparent inconsistency in the contribution of P2Y12 signaling to the overall platelet activation response, the present study investigated the possibility that the broad spectrum of inhibitory activity of this new generation of P2Y12 antagonists (i.e. MeSAMP and ARC69931MX) may derive from an elevation in platelet cAMP levels.Our data demonstrated that both 2MeSAMP and ARC69931MX do in fact significantly raise human platelet cAMP. Furthermore, this pharmacological effect is independent of P2Y12-Gi signaling and appears to proceed through activation of a separate Gs-coupled platelet receptor. Taken together, the results therefore indicate that these adenosine-based P2Y12 antagonists can produce their inhibition of platelet function through a cAMP-mediated mechanism. 相似文献
999.
The Family X DNA Polymerase from Deinococcus radiodurans Adopts
a Non-standard Extended Conformation
Nicolas Leulliot Lionel Cladière Fran?ois Lecointe Dominique Durand Ulrich Hübscher Herman van Tilbeurgh 《The Journal of biological chemistry》2009,284(18):11992-11999
Deinococcus radiodurans is an extraordinarily radioresistant
bacterium that is able to repair hundreds of radiation-induced double-stranded
DNA breaks. One of the players in this pathway is an X family DNA polymerase
(PolXDr). Deletion of PolXDr has been shown to decrease
the rate of repair of double-stranded DNA breaks and increase cell sensitivity
to gamma-rays. A 3′→5′ exonuclease activity that stops
cutting close to DNA loops has also been demonstrated. The present crystal
structure of PolXDr solved at 2.46-Å resolution reveals that
PolXDr has a novel extended conformation in stark contrast to the
closed “right hand” conformation commonly observed for DNA
polymerases. This extended conformation is stabilized by the C-terminal PHP
domain, whose putative nuclease active site is obstructed by its interaction
with the polymerase domain. The overall conformation and the presence of non
standard residues in the active site of the polymerase X domain makes
PolXDr the founding member of a novel class of polymerases involved
in DNA repair but whose detailed mode of action still remains enigmatic.DNA replication and repair are functions that are of vital importance for
the maintenance of cellular life. These functions are carried out by various
DNA replicating engines, most of them acting as multiprotein complexes.
Deinococcus radiodurans, a Gram-positive bacterium, is characterized
by an extraordinary resistance to ionizing radiation and desiccation. After
radiation induced cutting of its 3.28-megabase genome into hundreds of small
fragments, it is capable of reassembling it completely
(1). Different hypotheses have
been suggested to explain this radioresistance. A recently proposed mechanism
involves the creation of long linear DNA intermediates by an extended
synthesis-dependent strand annealing process, where overlapping chromosomal
fragments are used both as primers and as templates for synthesis of
complementary single strands
(2). Recircularization of
chromosomes would be assured by homologous recombination. Although DNA
polymerase I is one of the main enzymes involved in this process, it was shown
that other proteins affect double strand break repair efficiency in D.
radiodurans. One of these is an X family DNA polymerase
(PolXDr)5
(3). Cells devoid of
PolXDr protein show increased sensitivity to γ-irradiation
and a longer delay in the restoration of an intact genome after irradiation.
It was therefore proposed that PolXDr has an important role in
double strand break repair in D. radiodurans. The contribution of
PolXDr may become essential for instance when damage gets too
important or, alternatively, it may act in different repair pathways from
polymerase I. Indeed, some of the X DNA polymerases, such as Saccharomyces
cerevisiae Pol4 and human polymerase λ
(4) have been proposed to play
important roles in different DNA repair processes, including non-homologous
end-joining (5). It was shown
that PolXDr also has strong 3′→5′ exonuclease
activity that is stimulated by Mn2+
(6). This activity is
associated with proofreading mechanisms in other polymerase families and
encoded by protein domains or subunits distinct from the polymerase catalytic
domain (7). Curiously the
exonuclease activity of PolXDr is modulated upon encounter of a
stem-loop structure. The combination of both activities leads to the
hypothesis that PolXDr might be involved in DNA repair, potentially
non-homologous end-joining, by processing damaged DNA or repair intermediates,
thus generating substrates for other repair proteins
(6). Very recently an
orthologue of PolX from Bacillus subtilis was characterized. It was
shown that PolXBs is a template-directed DNA polymerase acting on
DNA gaps with a downstream 5′ phosphate group, suggesting it may play a
role in base excision repair
(8).DNA polymerases all combine a catalytic palm domain, a thumb domain,
binding double-stranded DNA, and a finger domain that fixes the incoming
nucleotide. The polymerase domain of the X family belongs to the
Polβ-like nucleotidyltransferase superfamily, sharing ∼25% amino acid
identity with the DNA polymerase domains of Polλ, Pol4, and Polβ.
PolXDr has a second domain at the C terminus called PHP, with
strong sequence identity with the histidinol phosphatase involved in histidine
transport in bacteria. Due to its similarity to histidinol phosphatase and the
presence of a trinuclear zinc site, the PolXDr PHP domain is
thought to function as phosphoesterase
(9). In the context of DNA
polymerases, this activity might be responsible for the degradation of
pyrophosphate, thus driving the polymerization reaction, or contributes to a
nuclease reaction that would be involved in proofreading the newly synthesized
strand. The deletion of the PHP domain also had a negative effect on survival
of γ-irradiated cells suggesting that this domain possesses a function
in DNA repair. Unexpectedly, deletion of the PHP domain destroys structure
modulated but not the general 3′→5′ exonuclease activity
(6). No activity could be
demonstrated for the PHP domain alone.In this report we present the crystal structure of PolXDr at
2.46-Å resolution. Surprisingly, PolXDr adopts a stretched
out conformation instead of the commonly observed closed right hand
conformation. In the active site of the polymerase catalytic domain, the two
universally conserved aspartates are replaced by two glutamates, whereas the
active site of the PHP domain is obstructed by its interaction with the
polymerase domain. 相似文献
1000.
Sana?MezghaniEmail author Olivier?Lesouhaitier Annabelle?Merieau Sylvie?Chevalier Nicole?Orange Marc?Feuilloley Lionel?Cazin 《Annals of microbiology》2009,59(2):379
In the present study the effect of infection on the resting membrane potential and whole-voltage-activated currents of neurons was determined usingPseudomonas fluorescens and lipopolysaccharide (LPS) extracted from the same species. Electrophysiological studies were performed on cerebellar granule neurons using the whole cell configuration of the patch clamp technique. The resting potential of cells (?63.7±2 mV) was significantly less negative (?46.0±4.7 mV) in cells showing adherent bacteria (106 CFU/ml). In addition, the whole-voltage currents triggered by voltage pulses were markedly reduced in neurons incubated withP. Fluorescens (mean 58%). Treatment with LPS (200 ng/ml) extracted fromP. Fluorescens provoke also a significant depolarisation of the membrane of the granule neurons and a severely alteration of whole-voltage currents. Analysis of the current-voltage relationships of the peak currents revealed the implication of potassium channels in the response of the cells toP. Fluorescens and its LPS. The present report provides the first data on the effect of a living bacterium on the membrane electrical activity of neurons and comforts the recent observations of the high cytotoxic potential ofP. fluorescens. 相似文献