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151.
λ-Glutamylcysteine synthetase in higher plants: catalytic properties and subcellular localization 总被引:1,自引:0,他引:1
λ-Glutamylcysteine synthetase activity (EC 6.3.2.2) was analysed in Sephacryl S-200 eluents of extracts from cell suspension
cultures ofNicotiana tabacum L. cv. Samsun by determination of λ-glutamylcysteine as its monobromobimane derivative. The enzyme has a relative molecular
mass (Mr) of 60000 and exhibits maximal activity at pH 8 (50% at pH 7.0 and pH9.0) and an absolute requirement for Mg2+. With 0.2mM Cd2+ or Zn2+, enzyme activity was reduced by 35% and 19%, respectively. Treatment with 5 mM dithioerythritol led to a heavy loss of activity
and to dissociation into subunits (Mr 34000). Buthionine sulfoximine andl-methionine-sulfoximine, known as potent inhibitors of λ-glutamylcysteine synthetase from mammalian cells, were found to be
effective inhibitors of the plant enzyme too. The apparent Km values forl-glutamate,l-cysteine, and α-aminobutyrate were, respectively, 10.4mM, 0.19 mM, and 6.36 mM. The enzyme was completely inhibited by glutathione
(Ki=0.42 mM). The data indicate that the rate of glutathione synthesis in vivo may be influenced substantially by the concentration
of cysteine and glutamate and may be further regulated by feedback inhibition of λ-glutamylcysteine synthetase by glutathione
itself. λ-Glutamylcysteine synthetase is, like glutathione synthetase, localized in chloroplasts as well as in the cytoplasm.
Chloroplasts fromPisum sativum L. isolated on a Percoll gradient contained about 72% of the λ-glutamylcysteine synthetase activity in leaf cells and 48%
of the total glutathione synthetase activity. In chloroplasts ofSpinacia oleracea L. about 61% of the total λ-glutamylcysteine synthetase activity of the cells were found and 58% of the total glutathione
synthetase activity. These results indicate that glutathione synthesis can take place in at least two compartments of the
plant cell.
Dedicated to Professor A. Prison on the occasion of his 80th birthday 相似文献
152.
153.
S. Romagnoli M. Maddaloni C. Livini M. Motto 《TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik》1990,80(6):769-775
Summary To provide an insight into the molecular basis of heterosis, we investigated gene expression in primary root tips of a heterotic maize hybrid (B73 × Mo17) and its parental lines (B73 and Mo17). This analysis was carried out (i) by differential plaque hybridization of a recombinant cDNA library made to poly(A) RNA isolated from B73 × Mo17 primary root tips, and (ii) by comparing with two-dimensional gel electrophoresis proteins synthesized in vitro in the rabbit reticulocyte system by poly(A) RNA isolated, at different stages of development, from the three genotypes. The results showed that there are sets of proteins and mRNAs that are differentially synthesized and expressed in the F1 primary root tips in comparison to the parental lines. Moreover, results from the survey of 21 major in-vitrosynthesized polypeptide variants, from mRNAs of primary root tips of the parental lines and their F1 hybrid, indicated that in seven instances hybrid proteins translated in vitro were more abundant or possibly new. In most of the remaining cases, hybrid spots were similar in intensity to the same protein produced by one of the two parental lines. 相似文献
154.
Christiane Gulat-Marnay rée Lafitte Jean-Michel Arrang Jean-Charles Schwartz 《Journal of neurochemistry》1989,52(1):248-254
The cholinergic modulation of histamine release and synthesis was studied in rat brain slices or synaptosomes labeled with L-[3H]histidine. Carbachol in increasing concentrations progressively reduced the K+-induced [3H]histamine release from cortical slices. Pirenzepine, a preferential M1-receptor antagonist, reversed the carbachol effect in an apparently competitive manner and with Ki values of 1-6 X 10(-8) M. 11-[(2-[(Diethylamino)methyl]-1-piperidinyl)acetyl]-5,11-dihydro-6H- pyrido[2,3-b][1,4]benzodiazepine-6-one (AF-DX 116), considered a preferential M2-receptor antagonist, reversed the carbachol effect with a mean Ki of approximately 2 X 10(-7) M. Oxotremorine behaved as a partial agonist in the modulation of histamine release. Neostigmine, an acetylcholinesterase inhibitor, inhibited the K+-induced release of [3H]histamine from cortical slices, and the effect was largely reversed by pirenzepine, an observation suggesting a modulation by endogenous acetylcholine. The effects of carbachol and pirenzepine were observed with slices of other brain regions known to contain histaminergic nerve terminals or perikarya, as well as with cortical synaptosomes. The two drugs also modified, in opposite directions, [3H]histamine formation in depolarized cortical slices. In vivo oxotremorine inhibited [3H]histamine formation in cerebral cortex, and this effect was reversed by scopolamine. When administered alone, scopolamine failed to enhance significantly the 3H- labeled amine formation, a finding suggesting that muscarinic receptors are not activated by endogenous acetylcholine released under basal conditions. It is concluded that muscarinic heteroreceptors, directly located on histaminergic nerve terminals, control release and synthesis of histamine in the brain. These receptors apparently belong to the broad M1-receptor category and may correspond to a receptor subclass displaying a rather high affinity for AF-DX 116. 相似文献
155.
U. Gartenbach-Scharrer S. Habib K. -H. Neumann 《Plant Cell, Tissue and Organ Culture》1990,22(1):27-35
Freshly isolated explants of the secondary phloem of carrot roots were exposed to 14C-leucine for various periods from t0—to 18 h and the 14C labelling of protein was studied by 2-dimensional PAGE followed by fluorograph. The labelling pattern of proteins indicated a sequential activation of synthesis of about 130 proteins during the 18 h experimental period prior to the onset of cell division activity.Abbreviations IAA
indole acetic acid
- 2iP
2-isopentenyladenine
- PVP
polyvinylpyrrolidone
- CBB
Coomassie brilliant blue
- RuBPCase
ribulosebisphosphate carboxylase
- LSC
liquid scintillation counter
- spec.act.
specific radioactivity
- u.l.
uniformly labelled 相似文献
156.
Thymocyte growth peptide (TGP) initiates DNA synthesis in immature thymocytes and has previously been characterized as an acidic peptide isolated from calf thymus. We now report the isolation of TGP from sheep thymus and show it to be a nonapeptide with a large N-terminal blocking moiety characterized by high UV absorbance. The amino acid composition is identical to FTS, consisting of 2 Gly, 2 Ser, 2 Glx, 1 Ala, 1 Lys, 1 Asx. In contrast to FTS, TGP is acidic with an apparent isoelectric point of 4.2 and a high UV absorbance at 270–280 nm. Reverse phase chromatography of TGP at an acidic pH results in a change of the molecule and the appearance of two new compounds TGP-A and TGP-B, both with less than 50% of the original TGP activity. Full activity could be restored by the addition of ZnCl2 to TGP-A. Both TGP-A and B have some amino acid composition and high UV absorbance as native TGP. We propose that TGP consists of a non-peptide moiety bound to the N-terminal of the nonapeptide Glu-Ala-Lys-Ser-Gln-Gly-Gly-Ser-Asn and that the active molecule is stabilized by Zn2+. 相似文献
157.
158.
Increased Neostriatal Tyrosine Hydroxylation During Stress: Role of Extracellular Dopamine and Excitatory Amino Acids 总被引:2,自引:0,他引:2
Abstract: We examined the regulation of neostriatal tyrosine hydroxylation during acute stress, testing the hypothesis that excitatory amino acids (EAAs) contribute to the stress-evoked increase in dopamine (DA) synthesis. Dialysis probes implanted into neostriatum permitted delivery of drugs and sampling of extracellular fluid. Rats were exposed to 30 min of intermittent tail shock during infusion of an inhibitor of aromatic amino acid decarboxylase (AAAD), NSD-1015 (100 µM), and DOPA was measured in the dialysate. Tail shock was applied beginning either 15 min after the onset of NSD-1015 treatment (the initial rate of DOPA accumulation) or 75 min after the onset of treatment (when DOPA had approached steady state). Tail shock increased the steady-state levels of extracellular DOPA in neostriatum (+40%). However, there was no change in the initial rate of DOPA accumulation unless animals also received the D2 receptor antagonist eticlopride (50 nM), in which case an increase was observed (+228%). The impact of tail shock on the steady-state level of DOPA was attenuated by the D2 agonist quinpirole (100 µM), or by 2-amino-5-phosphonovalerate (APV) (100 µM) or 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) (100 µM), EAA antagonists acting at NMDA or d ,l -α-amino-3-hydroxy-5-methyl-4-isoxazole-4-propionate (AMPA) receptors, respectively. These data suggest that acute stress normally has little effect on tyrosine hydroxylation in neostriatum due to the inhibitory influence of DA in the extracellular fluid. However, when that influence is absent (e.g., during extended inhibition of DOPA decarboxylation or blockade of DA receptors), stress increases tyrosine hydroxylation via EAAs acting on NMDA and AMPA receptors. Thus, EAAs released from corticostriatal projections may stimulate DA synthesis and thereby restore dopaminergic activity under conditions in which the availability of DA for release has been compromised. 相似文献
159.
Michael P. Robertson Matthew Levy Stanley L. Miller 《Journal of molecular evolution》1996,43(6):543-550
The reaction of guanidine hydrochloride with cyanoacetaldehyde gives high yields (40–85%) of 2,4-diaminopyrimidine under the concentrated conditions of a drying lagoon model of prebiotic synthesis, in contrast to the low yields previously obtained under more dilute conditions. The prebiotic source of cyanoacetaldehyde, cyanoacetylene, is produced from electric discharges under reducing conditions. The effect of pH and concentration of guanidine hydrochloride on the rate of synthesis and yield of diaminopyrimidine were investigated, as well as the hydrolysis of diaminopyrimidine to cytosine, isocytosine, and uracil. Thiourea also reacts with cyanoacetaldehyde to give 2-thiocytosine, but the pyrimidine yields are much lower than with guanidine hydrochloride or urea. Thiocytosine hydrolyzes to thiouracil and cytosine and then to uracil. This synthesis would have been a significant prebiotic source of 2-thiopyrimidines and 5-substituted derivatives of thiouracil, many of which occur in tRNA. The applicability of these results to the drying lagoon model of prebiotic synthesis was tested by dry-down experiments where dilute solutions of cyanoacetaldehyde, guanidine hydrochloride, and 0.5m NaCl were evaporated over varying periods of time. The yields of diaminopyrimidine varied from 1 to 7%. These results show that drying lagoons and beaches may have been major sites of prebiotic syntheses. 相似文献
160.
Stefan Evers Barbara Casadewall Murielle Charles Sylvie Dutka-Malen Marc Galimand Patrice Courvalin 《Journal of molecular evolution》1996,42(6):706-712
Thed-alanine:d-alanine-ligase-related enzymes can have three preferential substrate specificities. Usually, these enzymes synthesized-alanyl-d-alanine. In vancomycin-resistant Gram-positive bacteria, structurally related enzymes synthesized-alanyl-d-lactate or Dalanyl-d-serine. The sequence of internal fragments of eight structurald-alanine:d-alanine ligase genes from enterococci has been determined. Alignment of the deduced amino acid sequences with those of other
related enzymes from Gram-negative and Gram-positive bacteria revealed the presence of four distinct sequence patterns in
the putative substrate-binding sites, each correlating with specificity to a particular substrate (d-alanine:d-lactate ligases exhibited two patterns). Phylogenetic analysis showed different clusters. The enterococcal subtree was largely
superimposable on that derived from 16S rRNA sequences. In lactic acid bacteria, structural divergence due to differences
in substrate specificity was observed. Glycopeptide resistance proteins VanA and VanB, the VanC-type ligases, and Dd1A and
DdlB from enteric bacteria andHaemophilus influenzae constituted separate clusters.
Correspondence to: P. Courvalin 相似文献