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991.
Expression and purification of a small heat shock protein from the plant pathogen Xylella fastidiosa 总被引:2,自引:0,他引:2
Azzoni AR Tada SF Rosselli LK Paula DP Catani CF Sabino AA Barbosa JA Guimarães BG Eberlin MN Medrano FJ Souza AP 《Protein expression and purification》2004,33(2):297-303
The small heat shock proteins (smHSPs) belong to a family of proteins that function as molecular chaperones by preventing protein aggregation and are also known to contain a conserved region termed alpha-crystallin domain. Here, we report the expression, purification, and partial characterization of a novel smHSP (HSP17.9) from the phytopathogen Xylella fastidiosa, causal agent of the citrus variegated chlorosis (CVC). The gene was cloned into a pET32-Xa/LIC vector to over-express the protein coupled with fusion tags in Escherichia coli BL21(DE3). The expressed HSP17.9 was purified by immobilized metal affinity chromatography (IMAC) and had its identity determined by mass spectrometry (MALDI-TOF). The correct folding of the purified recombinant protein was verified by circular dichroism spectroscopy. Finally, the HSP17.9 protein also proved to efficiently prevent induced aggregation of insulin, strongly indicating a chaperone-like activity. 相似文献
992.
Coimbra DG Almeida AG Jùnior JB da Silva LA Pimentel BJ Gitaì DL Moreira LS Silva-Filho EA de Andrade TG 《The new microbiologica》2011,34(4):425-427
We describe a case of wound infection by multidrug-resistant Staphylococcus sciuri in a patient admitted to hospital for injuries in Agreste Alagoas, Brazil, identified through broad-spectrum PCR and sequencing of 16S rDNA gene. Due to its high resistance profile, the infection was characterized as methicillin-resistant Staphylococcus presenting sensitive only to vancomycin and chloramphenicol. The injury resulting from trauma associated with infection resulted in amputation of the infected limb. 相似文献
993.
Rafael D. Guariento Luciana S. Carneiro Adriano Caliman Reinaldo Luiz Bozelli Francisco A. Esteves 《Aquatic Ecology》2011,45(4):561-569
This study examines how nutrients and light affect the relationship between autotrophic biomass and non-autotrophic periphyton
organic matter in a tropical black water lake biofilm community. We hypothesized that there is no positive correlation between
autotrophic and non-autotrophic organic matter in the periphytic community of a black water humic lake, where non-algal components
of periphyton can rely on carbon sources external to the periphyton matrix and where nutrient availability is low. Second,
we sought to test our hypothesis that non-autotrophic periphyton organic matter will benefit from nutrient enhancement in
a lake where the availability of DOC is high. We performed a field experiment using in situ lake mesocosms to manipulate nutrient
concentrations and light availability in a 2 × 2 factorial design. Control treatments (no nutrient added) and nutrient treatments
(N + P) were compared in different light conditions: high light (near surface water) and low light (near bottom). No positive
correlation was found between autotrophic biomass and non-autotrophic periphyton organic matter, but a negative correlation
was observed in high nutrient and light conditions. The low C:P and N:P ratios revealed that the non-autotrophic organic matter
mostly comprised a heterotrophic microbial biofilm. High levels of light and nutrients together caused significant changes
in periphyton community properties. The non-autotrophic periphyton organic matter was negatively affected by nutrient addition,
whereas autotrophic biomass was positively affected, especially in high light conditions. Our results strongly suggest that
non-autotrophic periphyton organic matter in a humic lake is primarily comprised of a bacterial biofilm that directly competes
for nutrients with autotrophs in the periphytic community. We also observed no effect of nutrient addition on periphyton growing
in light-limited conditions. These results suggest that heterotrophic periphytic organisms might experience carbon limitation
despite the high availability, but usually low quality, of dissolved carbon in the water column of humic lakes. 相似文献
994.
Gentile Francesco Ficetola Matteo E. Siesa Raoul Manenti Luciana Bottoni Fiorenza De Bernardi Emilio Padoa‐Schioppa 《Diversity & distributions》2011,17(6):1141-1151
Aim Early assessment of the impact of invasive alien species is crucial to set up timely management, but often the impact is evident when it is too late for action. We evaluated relationships between the alien crayfish, Procambarus clarkii, the distribution of native amphibians, and the abundance of their larvae. We assessed whether considering measures of reproductive success provide a more prompt measure of impact than considering just species distribution. Location One hundred and twenty‐five wetlands in Northern Italy, in an area recently invaded by P. clarkii. Methods We surveyed wetlands to assess the presence of breeding activity of amphibians and the distribution of P. clarkii. We measured the abundance of amphibian larvae before metamorphosis through pipe sampling. We built models analysing the relationships between amphibian and crayfish distribution, while taking into account spatial autocorrelation and environmental features. Analyses were performed at both the species level (generalized linear models and spatial eigenvector mapping) and community level (constrained redundancy analysis). Results In terms of breeding site distribution, only two amphibians (Lissotriton vulgaris and Hyla intermedia) were negatively associated with P. clarkii, while the relationships between other the species and P. clarkii were positive or not significant. However, larval abundance for all amphibian species was negatively associated with the alien crayfish. Analyses performed at community and single species levels yielded consistent results. Main conclusions Procambarus clarkii impacts amphibians through different processes. Newts probably avoid invaded wetlands for breeding. Other species attempt breeding in wetlands with crayfish, but suffer very low success. Considering distribution data alone would not provide a correct picture of the impact of this alien species; measures of reproductive success may allow a more accurate assessment of the impact. 相似文献
995.
Mattos KA Lara FA Oliveira VG Rodrigues LS D'Avila H Melo RC Manso PP Sarno EN Bozza PT Pessolani MC 《Cellular microbiology》2011,13(2):259-273
The predilection of Mycobacterium leprae (ML) for Schwann cells (SCs) leads to peripheral neuropathy, a major concern in leprosy. Highly infected SCs in lepromatous leprosy nerves show a foamy, lipid-laden appearance; but the origin and nature of these lipids, as well as their role in leprosy, have remained unclear. The data presented show that ML has a pronounced effect on host-cell lipid homeostasis through regulation of lipid droplet (lipid bodies, LD) biogenesis and intracellular distribution. Electron microscopy and immunohistochemical analysis of lepromatous leprosy nerves for adipose differentiation-related protein expression, a classical LD marker, revealed accumulating LDs in close association to ML in infected SCs. The capacity of ML to induce LD formation was confirmed in in vitro studies with human SCs. Moreover, via confocal and live-cell analysis, it was found that LDs are promptly recruited to bacterial phagosomes and that this process depends on cytoskeletal reorganization and PI3K signalling. ML-induced LD biogenesis and recruitment were found to be independent of TLR2 bacterial sensing. Notably, LD recruitment impairment by cytoskeleton drugs decreased intracellular bacterial survival. Altogether, our data revealed SC lipid accumulation in ML-containing phagosomes, which may represent a fundamental aspect of bacterial pathogenesis in the nerve. 相似文献
996.
Claudia Trappetti Luciana Gualdi Lorenzo Di Meola Prashant Jain Cindy C Korir Paul Edmonds Francesco Iannelli Susanna Ricci Gianni Pozzi Marco R Oggioni 《BMC microbiology》2011,11(1):75
Background
Different models for biofilm in Streptococcus pneumoniae have been described in literature. To permit comparison of experimental data, we characterised the impact of the pneumococcal quorum-sensing competence system on biofilm formation in three models. For this scope, we used two microtiter and one continuous culture biofilm system. 相似文献997.
Determination of the phosphorylated metabolites of gemcitabine and of difluorodeoxyuridine by LCMSMS
Honeywell RJ Giovannetti E Peters GJ 《Nucleosides, nucleotides & nucleic acids》2011,30(12):1203-1213
Gemcitabine is an established chemotherapy agent in several solid tumors. Its mechanism of action has been theoretically established and this is supported with strong experimental evidence. However, certain aspects of the resistance mechanism for this agent remain elusive. We present a method of analysis using tandem liquid chromatography and mass spectrometry that provides a broader, yet more focused view of the action of gemcitabine and its primary metabolite, difluorodeoxyuridine in relation to the (deoxy) nucleoside and (deoxy) nucleotide pools in tumor cell lines. Alcoholic cytosole extracts were incubated with alkaline phosphatase reducing the nucleotide pools to their respective nucleosides. Determination of the nucleoside content by a sensitive LCMSMS method before and after incubation enables the calculation of the total amount of phosphorylation of each (deoxy) nucleoside in the cell. Incubation with clinically relevant levels of gemcitabine (dFdC) or difluorodeoxyuridine (dFdU) for 24 hours enabled the determination of the changes in the (deoxy) nucleotide pools in relation to chemotherapeutic and toxicological effects. Confirmation of the presence of dFdC phosphorylation is presented as well as direct evidence of dFdU phosphorylation after both dFdC and dFdU treatment. Differences in the nucleotide pools are presented after dFdC and dFdU incubation, indicating that dFdU might have more chemotherapeutic properties than previously believed. 相似文献
998.
Phytoplasmas, uncultivable phloem-limited phytopathogenic wall-less bacteria, represent a major threat to agriculture worldwide. They are transmitted in a persistent, propagative manner by phloem-sucking Hemipteran insects. Phytoplasma membrane proteins are in direct contact with hosts and are presumably involved in determining vector specificity. Such a role has been proposed for phytoplasma transmembrane proteins encoded by circular extrachromosomal elements, at least one of which is a plasmid. Little is known about the interactions between major phytoplasma antigenic membrane protein (Amp) and insect vector proteins. The aims of our work were to identify vector proteins interacting with Amp and to investigate their role in transmission specificity. In controlled transmission experiments, four Hemipteran species were identified as vectors of "Candidatus Phytoplasma asteris", the chrysanthemum yellows phytoplasmas (CYP) strain, and three others as non-vectors. Interactions between a labelled (recombinant) CYP Amp and insect proteins were analysed by far Western blots and affinity chromatography. Amp interacted specifically with a few proteins from vector species only. Among Amp-binding vector proteins, actin and both the α and β subunits of ATP synthase were identified by mass spectrometry and Western blots. Immunofluorescence confocal microscopy and Western blots of plasma membrane and mitochondrial fractions confirmed the localisation of ATP synthase, generally known as a mitochondrial protein, in plasma membranes of midgut and salivary gland cells in the vector Euscelidius variegatus. The vector-specific interaction between phytoplasma Amp and insect ATP synthase is demonstrated for the first time, and this work also supports the hypothesis that host actin is involved in the internalization and intracellular motility of phytoplasmas within their vectors. Phytoplasma Amp is hypothesized to play a crucial role in insect transmission specificity. 相似文献
999.
1000.
Vieira SA Alves LF Duarte-Neto PJ Martins SC Veiga LG Scaranello MA Picollo MC Camargo PB do Carmo JB Neto ES Santos FA Joly CA Martinelli LA 《Ecology and evolution》2011,1(3):421-434
We estimated carbon and nitrogen stocks in aboveground biomass (AGB) and belowground biomass (BGB) along an elevation range in forest sites located on the steep slopes of the Serra do Mar on the north coast of the State of São Paulo, southeast Brazil. In elevations of 100 m (lowland), 400 m (submontane), and 1000 m (montane) four 1-ha plots were established, and above- (live and dead) and belowground (live and dead) biomass were determined. Carbon and nitrogen concentrations in each compartment were determined and used to convert biomass into carbon and nitrogen stocks. The carbon aboveground stock (CAGB) varied along the elevation range from approximately 110 to 150 Mg·ha−1, and nitrogen aboveground stock (NAGB), varied from approximately 1.0 to 1.9 Mg·ha−1. The carbon belowground stock (CBGB) and the nitrogen belowground stock (NBGB) were significantly higher than the AGB and varied along the elevation range from approximately 200–300 Mg·ha−1, and from 14 to 20 Mg·ha−1, respectively. Finally, the total carbon stock (CTOTAL) varied from approximately 320 to 460 Mg·ha−1, and the nitrogen total stock (NTOTAL) from approximately 15 to 22 Mg·ha−1. Most of the carbon and nitrogen stocks were found belowground and not aboveground as normally found in lowland tropical forests. The above- and belowground stocks, and consequently, the total stocks of carbon and nitrogen increased significantly with elevation. As the soil and air temperature also decreased significantly with elevation, we found a significantly inverse relationship between carbon and nitrogen stocks and temperature. Using this inverse relationship, we made a first approach estimate that an increase of 1°C in soil temperature would decrease the carbon and nitrogen stocks in approximately 17 Mg·ha−1 and 1 Mg·ha−1 of carbon and nitrogen, respectively. 相似文献