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21.
Cloning of a negative transcription factor that binds to the upstream conserved region of Moloney murine leukemia virus. 总被引:45,自引:19,他引:26 下载免费PDF全文
J R Flanagan K G Becker D L Ennist S L Gleason P H Driggers B Z Levi E Appella K Ozato 《Molecular and cellular biology》1992,12(1):38-44
The long terminal repeat of Moloney murine leukemia virus (MuLV) contains the upstream conserved region (UCR). The UCR core sequence, CGCCATTTT, binds a ubiquitous nuclear factor and mediates negative regulation of MuLV promoter activity. We have isolated murine cDNA clones encoding a protein, referred to as UCRBP, that binds specifically to the UCR core sequence. Gel mobility shift assays demonstrate that the UCRBP fusion protein expressed in bacteria binds the UCR core with specificity identical to that of the UCR-binding factor in the nucleus of murine and human cells. Analysis of full-length UCRBP cDNA reveals that it has a putative zinc finger domain composed of four C2H2 zinc fingers of the GLI subgroup and an N-terminal region containing alternating charges, including a stretch of 12 histidine residues. The 2.4-kb UCRBP message is expressed in all cell lines examined (teratocarcinoma, B- and T-cell, macrophage, fibroblast, and myocyte), consistent with the ubiquitous expression of the UCR-binding factor. Transient transfection of an expressible UCRBP cDNA into fibroblasts results in down-regulation of MuLV promoter activity, in agreement with previous functional analysis of the UCR. Recently three groups have independently isolated human and mouse UCRBP. These studies show that UCRBP binds to various target motifs that are distinct from the UCR motif: the adeno-associated virus P5 promoter and elements in the immunoglobulin light- and heavy-chain genes, as well as elements in ribosomal protein genes. These results indicate that UCRBP has unusually diverse DNA-binding specificity and as such is likely to regulate expression of many different genes. 相似文献
22.
Y Ma B I Wilson S Bijvoet H E Henderson E Cramb G Roederer M R Ven Murthy P Julien H D Bakker J J Kastelein 《Genomics》1992,13(3):649-653
We have previously reported two common lipoprotein lipase (LPL) gene mutations underlying LPL deficiency in the majority of 37 French Canadians (Monsalve et al., 1990. J. Clin. Invest. 86: 728-734; Ma et al., 1991. N. Engl. J. Med. 324: 1761-1766). By examining the 10 coding exons of the LPL gene in another French Canadian patient, we have identified a third missense mutation that is found in two of the three remaining patients for whom mutations are undefined. This is a G to A transition in exon 6 that results in a substitution of asparagine for aspartic acid at residue 250. Using in vitro site-directed mutagenesis, we have confirmed that this mutation causes a catalytically defective LPL protein. In addition, the Asp250----Asn mutation was also found on the same haplotype in an LPL-deficient patient of Dutch ancestry, suggesting a common origin. This mutation alters a TaqI restriction site in exon 6 and will allow for rapid screening in patients with LPL deficiency. 相似文献
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25.
Peptide analogues compete with the binding of alpha-factor to its receptor in Saccharomyces cerevisiae 总被引:2,自引:0,他引:2
alpha-Factor, a secreted tridecapeptide pheromone, is required for mating between the a- and alpha-haploid mating types of Saccharomyces cerevisiae. An analogue of alpha-factor, [DHP8,DHP11,Nle12] tridecapeptide (where DHP represents 3,4-dehydro-L-proline and Nle represents norleucine), was catalytically reduced in the presence of 3H gas to produce a radiolabeled pheromone with high specific activity, purity, and biological activity. Association and dissociation kinetics indicated values of 4.9 x 10(4) M-1 s-1 for k1 and 1.1 x 10(-3) s-1 for k-1. Saturation binding studies gave an equilibrium dissociation constant equal to 2.3 x 10(-8) M, which approximated the kinetically derived KD of 2.2 x 10(-8) M. These values compare favorably to the previously determined KD of 6 x 10(-9) M (Jenness, D.D., Burkholder, A.C., and Hartwell, L.H. (1986) Mol. Cell. Biol. 6, 318-320). Scatchard analysis and dissociation in the presence of excess unlabeled ligand indicated interaction with a homogeneous population of noninteracting binding sites (13,000 sites/cell). A number of alpha-factor analogues, previously investigated for their structure-function relationships (Naider, F., and Becker, J.M. (1986) CRC Crit. Rev. Biochem. 21, 225-249), were used to compete with [3H]alpha-factor binding. Four tridecapeptides having conservative amino acid replacements bound strongly to the receptor. In contrast, [Phe3]alpha-factor and 10 des-Trp1-alpha-factor analogues bound to the receptor 1-3 orders of magnitude less effectively than did alpha-factor itself. The binding constants for all active pheromones correlated with biological activity. However, des-Trp1[Phe3]alpha-factor and des-Trp1-[Ala3]alpha-factor, which were not biologically active, still competed with alpha-factor binding, indicating that these analogues fail to induce a secondary signal necessary for biological response to the pheromone. One analogue, des-Trp1-[Cha3,L-Ala9]alpha-factor (where Cha represents cyclohexylalanine), was not biologically active and did not demonstrate binding to the receptor, whereas des-Trp1-[Cha3,D-Ala9]alpha-factor was active and bound to the receptor. This finding suggests that a type II beta-turn is necessary for binding of alpha-factor to its receptor and for subsequent biological activity. 相似文献
26.
Leukocyte stimulation: receptor, membrane, and metabolic events. Introduction and summary 总被引:5,自引:0,他引:5
E L Becker 《Federation proceedings》1986,45(7):2148-2150
27.
Analogues of the des-1-tryptophan,3-beta-cyclohexylalanine-alpha-factor of Saccharomyces cerevisiae, where the glycyl residue of position 9 was replaced by D-Ala, L-Ala, D-Leu, and L-Leu, were synthesized and evaluated by morphogenesis assays and circular dichroism spectroscopy. Synthesis was accomplished in solution phase with mixed anhydrides and p-nitrophenyl active esters as the coupling agents. All crude dodecapeptides were purified to greater than 98% homogeneity by preparative high-performance liquid chromatography on a reversed-phase column. The Gly9, D-Ala9, and D-Leu9 analogues elicited morphogenic alterations in MATa strains of S. cerevisiae at concentrations of 1-2 micrograms/mL and exhibited similar CD patterns in both trifluoroethanol and tris(hydroxymethyl)aminomethane buffer, pH 7.4. In contrast, the L-Ala9 and L-Leu9 analogues were more than 200 times less active in the morphogenesis assay and had markedly different CD spectra. These results demonstrate that the position 9 residue plays an important role in determining the biological activity and solution conformation of alpha-factor. We suggest the presence of a type II beta-turn in the Lys7-Gln10 region when the alpha-factor assumes its biologically active conformation. 相似文献
28.
We have cloned a cDNA coding for the smallest rat neurofilament protein. The cDNA is 861 nucleotides long coding for 287 amino acids from the internal alpha-helical region and the carboxy-terminal tail domain of the neurofilament protein. Comparison of the porcine, mouse and rat neurofilament protein sequences shows that the protein is highly conserved (greater than 93% identity). Blot analysis indicates that the cDNA is derived from a single neurofilament gene that codes for two different poly(A)+ mRNA species. 相似文献
29.
The inhibition of neutrophil granule enzyme secretion and chemotaxis by pertussis toxin 总被引:21,自引:2,他引:19 下载免费PDF全文
E L Becker J C Kermode P H Naccache R Yassin M L Marsh J J Munoz R I Sha'afi 《The Journal of cell biology》1985,100(5):1641-1646
Pertussis toxin treatment of rabbit peritoneal neutrophils causes a concentration-dependent inhibition of granule enzyme secretion induced by formylmethionyl-leucyl-phenylalanine, C5a, and leukotriene B4. It also inhibits chemotaxis induced by formylmethionyl-leucyl-phenylalanine. The same toxin treatment, however, has no effect on granule enzyme secretion induced by the calcium ionophore A23187 or phorbol 12-myristate 13-acetate. Moreover, pertussis toxin treatment does not affect either the number or affinity of the formylpeptide receptors on the neutrophil nor does it have any effect on the unstimulated levels of cyclic AMP (cAMP) or the transient rise in cAMP induced by chemotactic factor stimulation in these cells. We hypothesize that pertussis toxin, as in other cells, interacts with a GTP binding regulatory protein identical with or analogous to either Ni or transducin which mediates the receptor-induced inhibition or activation of a target protein or proteins required in neutrophil activation. The nature of the target protein is unknown, but it is not the catalytic unit of adenylate cyclase. The target protein acts after binding of chemotactic factor to its receptor in the sequence that leads to the receptor-induced rise in intracellular Ca2+. It does not affect the responses elicited by the direct introduction of calcium into the cells or the activity of protein kinase C. 相似文献
30.
Zusammenfassung Regionale und artliche Unterschiede der Rückstände von Quecksilber in Eiern von sieben Brutvogelarten der deutschen Nordseeküste wurden untersucht. Bei vier Arten stellten wir deutliche geographische Unterschiede in der Kontamination fest (Abb. 1). Höchstgehalte fanden wir in Eiern von der Elbmündung (Region V). Austernfischer- und Flußseeschwalbeneier von der inneren Deutschen Bucht (Region VI) enthielten größere Mengen als solche von Nordfriesland (Region VII). Der gesetzliche Richtwert für Eier (0,03 mg/kg) wurde von 353 der 355 untersuchten Eier überschritten. Fluß- und Brandseeschwalbe waren am stärksten, Brandgans, Austernfischer und Lachmöwe am wenigsten mit Quecksilber kontaminiert (Abb. 2, 4.2.). Mit Ausnahme von Brandgans und Silbermöwe erreichten die Rückstände in vielen Eiern der anderen Arten, an der Elbe sogar in allen Eiern, eine den Bruterfolg gefährdende Größenordnung, insbesondere bei der Flußseeschwalbe.
Pollutants in eggs of birds breeding at the German North Sea Coast. II. Mercury
Summary Regional and interspecific differences of mercury residues were investigated in the eggs of seven species breeding at the German North Sea Coast (Tadorna tadorna, Haematopus ostralegus, Charadrius hiaticula, Larus argentatus, L. ridibundus, Sterna hirundo, S. sandvicensis). We found distinct geographical differences in contamination (Fig. 1): In eggs from the Elbe Eastuary (region V) maximum residues of mercury were ascertained. Eggs of Oystercatcher and Common Tern from the Bay of Helgoland (region VI) contained larger quantities than eggs from Northern Frisia (region VII). 353 from 355 eggs examined showed a value exceeding the legal standard value for eggs which is 0,03 mg/kg in Germany. Common Tern and Sandwich Tern eggs had the highest, Shelduck, Oystercatcher and Blackheaded Gull eggs the lowest mercury contamination (Fig. 2, 4.2.). With the exception of Shelduck and Herring Gull, residues in many eggs of the other species — at the river Elbe in all eggs — came up to a range endangering breeding success, especially in Common Terns.相似文献