全文获取类型
收费全文 | 22854篇 |
免费 | 1836篇 |
国内免费 | 11篇 |
出版年
2023年 | 91篇 |
2022年 | 207篇 |
2021年 | 393篇 |
2020年 | 239篇 |
2019年 | 339篇 |
2018年 | 399篇 |
2017年 | 356篇 |
2016年 | 584篇 |
2015年 | 1007篇 |
2014年 | 1150篇 |
2013年 | 1435篇 |
2012年 | 1634篇 |
2011年 | 1552篇 |
2010年 | 1011篇 |
2009年 | 940篇 |
2008年 | 1230篇 |
2007年 | 1168篇 |
2006年 | 1173篇 |
2005年 | 1139篇 |
2004年 | 1054篇 |
2003年 | 937篇 |
2002年 | 981篇 |
2001年 | 327篇 |
2000年 | 282篇 |
1999年 | 313篇 |
1998年 | 305篇 |
1997年 | 238篇 |
1996年 | 255篇 |
1995年 | 227篇 |
1994年 | 232篇 |
1993年 | 227篇 |
1992年 | 225篇 |
1991年 | 193篇 |
1990年 | 167篇 |
1989年 | 191篇 |
1988年 | 175篇 |
1987年 | 142篇 |
1986年 | 134篇 |
1985年 | 151篇 |
1984年 | 177篇 |
1983年 | 149篇 |
1982年 | 165篇 |
1981年 | 123篇 |
1980年 | 122篇 |
1979年 | 120篇 |
1978年 | 94篇 |
1977年 | 102篇 |
1976年 | 67篇 |
1974年 | 72篇 |
1973年 | 74篇 |
排序方式: 共有10000条查询结果,搜索用时 15 毫秒
111.
Tjakko Abee Jan Knol Klaas J. Hellingwerf Evert P. Bakker Annette Siebers Wil N. Konings 《Archives of microbiology》1992,158(5):374-380
Cells of the purple non-sulphur bacterium Rhodobacter sphaeroides express a high-affinity K+ uptake system when grown in media with low K+ concentrations. Antibodies againts the catalytic KdpB protein or the whole KdpABC complex of Escherichia coli crossreact with a 70.0 kDa R. sphaeroides protein that was expressed only in cells grown in media with low K+ concentrations. In membranes derived from R. sphaeroides cells grown with low K+ concentrations (induced cells), a high ATPase activity could be detected when assayed in Tris-HCl pH 8.0 containing 1 mM MgSO4. This ATPase activity increased upon addition of 1 mM KCl from 166 to 289 mol ATP hydrolysed x min-1 x g protein-1 (1.7-fold stimulation). The K+-stimulated ATPase activity was inhibited approximately 93% by 0.5 mM vanadate but hardly by N,N-dicyclohexylcarbo-diimide (DCCD). These results indicate that the inducible K+-ATPase in R. sphaeroides resembles the Kdp K+-translocating ATPase of Escherichia coli. This Kdp-like transport system is also expressed in R. capsulatus and Rhodospirillum rubrum during growth in media with low K+ concentrations suggesting a wide distribution of this transport system among phototrophic bacteria.Abbreviations
electrical potential difference across the cytoplasmic membrane
- pH
pH difference across the cytoplasmic membrane
- BSA
bovine serum albumine
- PAGE
polyacrylamide gel electrophoresis
- HEPES
4-(2-hydroxyethyl)-1-piperazine-ethanesulfonic acid
- PMSF
phenyl-methyl-sulfonyl fluoride
- DCCD
N,N-dicyclohexylcarbodiimide
- AIB
2--aminoisobutyric acid
- TMG
methyl--d-thiogalactopyranoside 相似文献
112.
The velocities of single stranded DNA molecules in denaturing polyacrylamide gels during symmetric and asymmetric field inversion were measured at different pulse times and gel concentrations. Under the conditions chosen in our study, pulse times as short as a few milliseconds lead to a retardation of DNA molecules larger than 400 bases. We found that a field inversion with an electric field in the forward direction of about double the strength of that applied in the backward direction is a good compromise between the degree of retardation, the temperature control requirements and the run time of the gel. 相似文献
113.
Influence of soil type on the transfer of plasmid RP4p from Pseudomonas fluorescens to introduced recipient and to indigenous bacteria 总被引:1,自引:0,他引:1
Agnes Richaume Eric Smit Geneviève Faurie Jan D. van Elsas 《FEMS microbiology letters》1992,101(4):281-291
Abstract Transfer of plasmid RP4p from introduced Pseudomonas fluorescens to a co-introduced recipient strain or to members of the indigenous bacterial population was studied in four different soils of varying texture planted with wheat. Donor and recipient strains showed good survival in the four soils throughout the experiment. The numbers of transconjugants found in donor and recipient experiments in two soils, Ede loamy sand and Löss silt loam were significantly higher in the rhizosphere than in corresponding bulk soil. In the remaining two soils, Montrond and Flevo silt loam, transconjugant numbers were not significantly higher in the rhizosphere than in the bulk soil.
The combined utilization of a specific bacteriophage eliminate the donor strain and the pat sequence as a specific marker to detect RP4p was found to be very efficient in detecting indigenous transconjugants under various environmental conditions. The numbers of indigenous transconjugants were consistently higher in rhizosphere than bull soil. A significant rhizosphere effect on transconjugant numbers of transconjugants were recovered from Flevo and Montrond silt loam; these soils possess characteristics such as clay or organic matter contents which may be favorable to conjugation. 相似文献
The combined utilization of a specific bacteriophage eliminate the donor strain and the pat sequence as a specific marker to detect RP4p was found to be very efficient in detecting indigenous transconjugants under various environmental conditions. The numbers of indigenous transconjugants were consistently higher in rhizosphere than bull soil. A significant rhizosphere effect on transconjugant numbers of transconjugants were recovered from Flevo and Montrond silt loam; these soils possess characteristics such as clay or organic matter contents which may be favorable to conjugation. 相似文献
114.
Coen van den Berg Ferdinand A. Sibbing Jan W. M. Osse Wim Hoogenboezem 《Environmental Biology of Fishes》1992,33(1-2):105-124
Synopsis The filter feeding organ of cyprinid fishes is the branchial sieve, which consists of a mesh formed by gill rakers and tiny channels on the gill arches. In order to establish its possible role during growth we measured the following morphological gill raker parameters over a range of sizes in three cyprinid fishes, bream, white bream and roach: inter raker distance, bony raker length, raker width, cushion length and channel width. At any given standard length common bream has the largest inter raker distance, roach the lowest and white bream is intermediate. In the comb model of filter feeding the inter raker distance is considered to be a direct measure of the mesh size and retention ability (= minimal size of prey that can be retained) of a filter. For the three species under study there is a conflict between the comb model and experimental data on particle retention. Lammens et al. (1987) found that common bream has a large retention ability whereas roach and white bream have a much smaller one. A new model, the channel model (Hoogenboezem et al. 1991) has been developed for common bream; in this model the lateral gill rakers can regulate the mesh size of the medial channels on the other side of the gill slit. The present data indicate that this model is not appropriate for white bream and roach. At any given standard length white bream and roach only reach 70% of the raker length of common bream, which means that in this model the gill slits should to be very narrow during filter feeding. The gill rakers consist of a bony raker and a fleshy cushion. The bony rakers have a rather long needle-like part outside the cushion in bream, but not in white bream and roach which have blunt gill rakers. Blunt gill rakers are not suited to reduce the diameter of the medial channels. The comb model seems more appropriate for white bream and roach, but doubts about the validity of this simple model remain. The sum of the areas of the medial channels is an approximation of the area through which water flows in the filter. This channel area therefore gives an impression of the capacity or flow rate of the filter. With this capacity estimation and an estimation of energy consumption we calculated an energy ratio of filter feeding. The energy ratio decreases with increasing standard length with an exponent close to the expected exponent of -0.40. The energy ratio is highest in bream, intermediate in white bream and lowest in roach. 相似文献
115.
The response of a bumblebee goby,Brachygobius sabanus,to chemical stimuli from injured conspecifics 总被引:1,自引:0,他引:1
Synopsis
Brachygobius sabanus move less often and spend less time swimming when they detect chemicals released from injured conspecifics. This resembles the alarm response found in ostariophysan fishes, darters, and at least one other gobiid. Chemicals from injured Poecilia reticulata do not induce an alarm response in B. sabanus. 相似文献
116.
Differential expression of K+ channel mRNAs in the rat brain and down-regulation in the hippocampus following seizures. 总被引:4,自引:0,他引:4
K+ channels are major determinants of membrane excitability. Differences in neuronal excitability within the nervous system may arise from differential expression of K+ channel genes, regulated spatially in a cell type-specific manner, or temporally in response to neuronal activity. We have compared the distribution of mRNAs of three K+ channel genes, Kv1.1, Kv1.2, and Kv4.2 in rat brain, and examined activity-dependent changes following treatment with the convulsant drug pentylenetetrazole. Both regional and cell type-specific differences of K+ channel gene expression were found. In addition, seizure activity caused a reduction of Kv1.2 and Kv4.2 mRNAs in the dentate granule cells of the hippocampus, raising the possibility that K+ channel gene regulation may play a role in long-term neuronal plasticity. 相似文献
117.
Eyvind J. Paulssen Ruth H. Paulssen Kaare M. Gautvik Jan O. Gordeladze 《Cellular signalling》1992,4(6):747-755
We have investigated the possibility that adenylyl cyclase (AC) activity and membrane protein levels of the -subunits of the stimulatory and inhibitory G-proteins of AC (Gs and Gi−2) in cultured prolactin-producing rat pituitary adenoma cells (GH3 cells) are modulated by phospholipase C (PLC)-generated second messengers. Pretreatment of cells (6–48 h) with ionomycin (1 μM) or 1-oleoyl-2-acetylglycerol (OAG; 1μM) showed that ionomycin regulated Gs levels in a time-dependent, biphasic manner; a two-fold increase followed a 40% initial reduction, while OAG lowered Gs levels by more than 50% at all time-points. Gi−2 levels remained unchanged by both pretreatments. OAG, but not ionomycin, increased basal AC activity without increasing enzyme protein levels. Alterations in AC responsiveness to peptide hormones (e.g. thyroliberin and vasoactive intestinal peptide) correlated to membrane Gs protein -subunit content. These results demonstrate the involvement of G-protein translation regulation as one mechanism of ‘cross-talk’ between the PLC- and AC-dependent signalling pathways. 相似文献
118.
Viability measurements of hybridoma cells in suspension cultures 总被引:1,自引:0,他引:1
José M. Coco-Martin Jan W. Oberink Tiny A. M. van der Velden-de Groot E. Coen Beuvery 《Cytotechnology》1992,8(1):57-64
Several methods were applied to determine the viability of hybridoma cells in suspension. These methods include dye inclusion
and exclusion assays such as the classical trypan blue exclusion assay, the propidium iodide (PI) exclusion assay and the
fluorescein diacetate (FDA) inclusion assay. Furthermore, the relation was studied between release of lactate dehydrogenase
(LDH) by hybridoma cells and their viability. Also the ATP content of the cells and cellular heterogeneity as measured with
a flow cytometer were determined in relation to cellular viability.
The dye inclusion and exclusion assays using trypan blue, FDA, PI were shown to be useful methods to determine cellular viability.
With the FDA and PI methods it was possible to obtain additional information about cells which are in a transition state between
viable and non-viable. The viability according to the scatter properties of the cells appears to reflect the overall condition
of the cells, although interpretation of the results is difficult. Measurement of LDH release in the culture fluid or the
cytoplasmic ATP content could not be used as parameters for cell viability. 相似文献
119.
Interaction of a cellular 57-kilodalton protein with the internal translation initiation site of foot-and-mouth disease virus. 总被引:21,自引:10,他引:11
A cellular 57-kDa protein (p57) that binds specifically to the internal translation initiation site in the 5' untranslated region of foot-and-mouth disease virus RNA was detected in cell extracts of different mammalian species by UV cross-linking. The protein binds to two distinct sites of the translation control region which have as the only common sequence a UUUC motif. The first binding site consists of a conserved hairpin structure, whereas the second binding site contains an essential pyrimidine-rich region without obvious secondary structure. Competition experiments indicate that the complexes with the two binding sites were formed by a single p57 species. The protein binds also to the 5' untranslated region of other picornaviruses. Results from footprint analyses with foot-and-mouth disease RNA suggest the participation of additional cellular factors in the translation initiation complex. 相似文献
120.
Signal peptidase I overproduction results in increased efficiencies of export and maturation of hybrid secretory proteins in Escherichia coli 总被引:4,自引:0,他引:4
Jan Maarten van Dijl Anne de Jong Hilde Smith Sierd Bron Gerard Venema 《Molecular & general genetics : MGG》1991,227(1):40-48
Summary The effects of 25-fold overproduction ofEscherichia coli signal peptidase I (SPase I) on the processing kinetics of various (hybrid) secretory proteins, comprising fusions between
signal sequence functions selected from theBacillus subtilis chromosome and the mature part of TEM-β-lactamase, were studied inE. coli. One precursor (pre[A2d]-β-lactamase) showed an enhanced processing rate, and consequently, a highly improved release of
the mature enzyme into the periplasm. A minor fraction of a second hybrid precursor (pre[Al3i]-β-lactamase), which was not
processed under standard conditions of SPase I synthesis, was shown to be processed under conditions of SPase I overproduction.
However, this did not result in efficient release of the mature β-lactamase into the periplasm. In contrast, the processing
rates of wild-type pre-β-lactamase and pre(A2)-β-lactamase, already high under standard conditions, were not detectably altered
by SPase I overproduction. These results demonstrate that the availability of SPase I can be a limiting factor in protein
export inE. coli, in particular with respect to (hybrid) precursor proteins showing low (SPase I) processing efficiencies. 相似文献