全文获取类型
收费全文 | 2416篇 |
免费 | 110篇 |
国内免费 | 73篇 |
出版年
2023年 | 21篇 |
2022年 | 24篇 |
2021年 | 43篇 |
2020年 | 47篇 |
2019年 | 70篇 |
2018年 | 68篇 |
2017年 | 41篇 |
2016年 | 48篇 |
2015年 | 100篇 |
2014年 | 198篇 |
2013年 | 224篇 |
2012年 | 173篇 |
2011年 | 178篇 |
2010年 | 114篇 |
2009年 | 122篇 |
2008年 | 85篇 |
2007年 | 131篇 |
2006年 | 90篇 |
2005年 | 97篇 |
2004年 | 67篇 |
2003年 | 68篇 |
2002年 | 58篇 |
2001年 | 45篇 |
2000年 | 32篇 |
1999年 | 36篇 |
1998年 | 27篇 |
1997年 | 30篇 |
1996年 | 21篇 |
1995年 | 35篇 |
1994年 | 43篇 |
1993年 | 31篇 |
1992年 | 24篇 |
1991年 | 26篇 |
1990年 | 24篇 |
1989年 | 13篇 |
1988年 | 17篇 |
1987年 | 12篇 |
1986年 | 10篇 |
1985年 | 13篇 |
1984年 | 15篇 |
1983年 | 12篇 |
1982年 | 7篇 |
1981年 | 8篇 |
1980年 | 7篇 |
1979年 | 10篇 |
1978年 | 6篇 |
1977年 | 5篇 |
1976年 | 10篇 |
1975年 | 4篇 |
1970年 | 2篇 |
排序方式: 共有2599条查询结果,搜索用时 15 毫秒
1.
《Cryobiology》2020
Cryopreservation, the freezing and later warming of biological samples with minimal loss of viability, is important in many scientific disciplines. For some applications, particularly those where there is limited available material, it is critical to ensure the maximal survival rates of cryopreserved materials. Most of the challenges encountered with such techniques take place after the warming process where cryodamage affects cell viability and future development. Here we have used the nematode Caenorhabditis elegans to investigate the effects of cryodamage caused by slow-freezing. We find that freezing results in the death of some worms, with an approximately 40% reduction in the number of worms that develop in the frozen populations, but that the effects on worms that survive are limited. For example, there are no differences in the lifetime fecundity or in lifespan between frozen and control worms, although early fecundity and body size was reduced in frozen worms. Similarly, analyses of body wall muscle structure and of pharyngeal function indicates that muscle development and function are not significantly affected by freezing. We do however determine that freezing increases the rates of matricidal hatching, where progeny hatch within the mother. Overall, these results indicate that, for worms that survive, cryopreservation produces limited long-term effects, but do indicate that some phenotypes could be used in further analyses of the cellular damage induced by cryopreservation. 相似文献
2.
Jiaohong Zhao Fudan Gao Jingsong Zhang Teruo Ogawa Weimin Ma 《The Journal of biological chemistry》2014,289(39):26669-26676
Two mutants that grew faster than the wild-type (WT) strain under high light conditions were isolated from Synechocystis sp. strain PCC 6803 transformed with a transposon-bearing library. Both mutants had a tag in ssl1690 encoding NdhO. Deletion of ndhO increased the activity of NADPH dehydrogenase (NDH-1)-dependent cyclic electron transport around photosystem I (NDH-CET), while overexpression decreased the activity. Although deletion and overexpression of ndhO did not have significant effects on the amount of other subunits such as NdhH, NdhI, NdhK, and NdhM in the cells, the amount of these subunits in the medium size NDH-1 (NDH-1M) complex was higher in the ndhO-deletion mutant and much lower in the overexpression strain than in the WT. NdhO strongly interacts with NdhI and NdhK but not with other subunits. NdhI interacts with NdhK and the interaction was blocked by NdhO. The blocking may destabilize the NDH-1M complex and repress the NDH-CET activity. When cells were transferred from growth light to high light, the amounts of NdhI and NdhK increased without significant change in the amount of NdhO, thus decreasing the relative amount of NdhO. This might have decreased the blocking, thereby stabilizing the NDH-1M complex and increasing the NDH-CET activity under high light conditions. 相似文献
3.
Abstract Four isolates of the non-motile species Aeromonas media , icluding the type strain, were demonstrated by transmission electronk microcopy to possess polar flagella. Motility appeared to be restricted to old, regularly subcultured strains, and coincided with an increase in colony size and a concomitant reduction in the amount of brown diffusible pigment produced by colonies on tryptone soya agar. 相似文献
4.
5.
Tzong-Yuan Lin Tobias Werther Jae-Hun Jeoung Holger Dobbek 《The Journal of biological chemistry》2012,287(45):38338-38346
The three-component toluene dioxygenase system consists of an FAD-containing reductase, a Rieske-type [2Fe-2S] ferredoxin, and a Rieske-type dioxygenase. The task of the FAD-containing reductase is to shuttle electrons from NADH to the ferredoxin, a reaction the enzyme has to catalyze in the presence of dioxygen. We investigated the kinetics of the reductase in the reductive and oxidative half-reaction and detected a stable charge transfer complex between the reduced reductase and NAD+ at the end of the reductive half-reaction, which is substantially less reactive toward dioxygen than the reduced reductase in the absence of NAD+. A plausible reason for the low reactivity toward dioxygen is revealed by the crystal structure of the complex between NAD+ and reduced reductase, which shows that the nicotinamide ring and the protein matrix shield the reactive C4a position of the isoalloxazine ring and force the tricycle into an atypical planar conformation, both factors disfavoring the reaction of the reduced flavin with dioxygen. A rapid electron transfer from the charge transfer complex to electron acceptors further reduces the risk of unwanted side reactions, and the crystal structure of a complex between the reductase and its cognate ferredoxin shows a short distance between the electron-donating and -accepting cofactors. Attraction between the two proteins is likely mediated by opposite charges at one large patch of the complex interface. The stability, specificity, and reactivity of the observed charge transfer and electron transfer complexes are thought to prevent the reaction of reductaseTOL with dioxygen and thus present a solution toward conflicting requirements. 相似文献
6.
《Molecular & cellular proteomics : MCP》2022,21(12):100438
Human pancreatic stellate cells (HPSCs) are an essential stromal component and mediators of pancreatic ductal adenocarcinoma (PDAC) progression. Small extracellular vesicles (sEVs) are membrane-enclosed nanoparticles involved in cell-to-cell communications and are released from stromal cells within PDAC. A detailed comparison of sEVs from normal pancreatic stellate cells (HPaStec) and from PDAC-associated stellate cells (HPSCs) remains a gap in our current knowledge regarding stellate cells and PDAC. We hypothesized there would be differences in sEVs secretion and protein expression that might contribute to PDAC biology. To test this hypothesis, we isolated sEVs using ultracentrifugation followed by characterization by electron microscopy and Nanoparticle Tracking Analysis. We report here our initial observations. First, HPSC cells derived from PDAC tumors secrete a higher volume of sEVs when compared to normal pancreatic stellate cells (HPaStec). Although our data revealed that both normal and tumor-derived sEVs demonstrated no significant biological effect on cancer cells, we observed efficient uptake of sEVs by both normal and cancer epithelial cells. Additionally, intact membrane-associated proteins on sEVs were essential for efficient uptake. We then compared sEV proteins isolated from HPSCs and HPaStecs cells using liquid chromatography–tandem mass spectrometry. Most of the 1481 protein groups identified were shared with the exosome database, ExoCarta. Eighty-seven protein groups were differentially expressed (selected by 2-fold difference and adjusted p value ≤0.05) between HPSC and HPaStec sEVs. Of note, HPSC sEVs contained dramatically more CSE1L (chromosome segregation 1–like protein), a described marker of poor prognosis in patients with pancreatic cancer. Based on our results, we have demonstrated unique populations of sEVs originating from stromal cells with PDAC and suggest that these are significant to cancer biology. Further studies should be undertaken to gain a deeper understanding that could drive novel therapy. 相似文献
7.
A simple and convenient method has been tested for the steriltzation of nutrient media for long-term cultivation of plant cells. Diethylpyrocarbonate is suitable for this task in concentrations about 1000 mg l-1 The cells cultivated for 15 subsequent passages on media treated by DPC had the same growth parameters, production pattern and ability to transform exogeneous organic compounds as did the controls. The method is suitable for the preparatian of both liquid and agar media, for stabilization of stock solutions and for sterilization of cultivation vessels and tubing.Abbreviations DPC
diethylpyrocarbonate
- medium MS
nutrient medium according to Murashige and Skoog
- NAA
naphtaleneacetic acid 相似文献
8.
Thierry Brun Jean Péduzzi Manuela Marin Caniça Gérard Paul Pierre Névot Michel Barthélémy Roger Labia 《FEMS microbiology letters》1994,120(1-2):111-117
Abstract A novel procedure was used to purify a cytosolic chitinase from Candida albicans to electrophoretic homogeneity. The results represent the first demonstration of the purification of a fungal intracellular chitinase using the criterion of a single band detected following silver-staining of a polyacrylamide gel run under denaturing conditions. Purified chitinase had pH and temperature optima of 5.0 and 50°C, respectively. Inhibition of enzyme activity by allosamidin was pH-dependent occuring maximally at pH 8.0. Phospholipids had similar marked and highly specific effects on the activities of both the purified soluble enzyme and a solubilized microsomal chitinase from C. albicans . Evidence is provided for the existence of a complex chitinolytic system in this organism. 相似文献
9.
Insufficient oxygen transfer in a highly viscous media with varying viscosity occurs during the degradation of starch byBacillus licheniformis. Oxygen transfer rate (OTR) decreased below 0.3 gl–1 h–1 for viscosities above 5 to 10 mPa.s and agitation speeds lower than 710 rev/min. Increasing agitation speeds by 30 to 50% compensated for the decrease in OTR for viscosities between 10 and 200 mPa.s. A dual impeller is considered essential for growth improvement. 相似文献
10.
Respiratory complex I couples the transfer of electrons from NADH to ubiquinone and the translocation of protons across the mitochondrial membrane. A detailed understanding of the midpoint reduction potentials (Em) of each redox center and the factors which influence those potentials are critical in the elucidation of the mechanism of electron transfer in this enzyme. We present accurate electrostatic interaction energies for the iron-sulfur (FeS) clusters of complex I to facilitate the development of models and the interpretation of experiments in connection to electron transfer (ET) in this enzyme. To calculate redox titration curves for the FeS clusters it is necessary to include interactions between clusters, which in turn can be used to refine Em values and validate spectroscopic assignments of each cluster. Calculated titration curves for clusters N4, N5, and N6a are discussed. Furthermore, we present some initial findings on the electrostatics of the redox centers of complex I under the influence of externally applied membrane potentials. A means of determining the location of the FeS cofactors within the holo-complex based on electrostatic arguments is proposed. A simple electrostatic model of the protein/membrane system is examined to illustrate the viability of our hypothesis. 相似文献