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191.
Whitehouse DB; Tomkins J; Lovegrove JU; Hopkinson DA; McMillan WO 《Molecular biology and evolution》1998,15(4):456-462
The expanding molecular database provides unparalleled opportunities for
characterizing genes and for studying groups of related genes. We use
sequences drawn from the database to construct an evolutionary framework
for examining the important glycolytic enzyme phosphoglucomutase (PGM).
Phosphoglucomutase plays a pivotal role in the synthesis and utilization of
glycogen and is present in all organisms. In humans, there are three
well-described isozymes, PGMI, PGM2, and PGM3. PGM1 was cloned 5 years ago;
however, repeated attempts using both immunological approaches and
molecular probes designed from PGM1 have failed to isolate either PGM2 or
PGM3. Using a phylogenetic strategy, we first identified 47 highly
divergent prokaryotic and eukaryotic PGM-like sequences from the database.
Although overall amino acid identity often fell below 20%, the relative
order, position, and sequence of three structural motifs, the active site
and the magnesium-- and sugar-binding sites, were conserved in all 47
sequences. The phylogenetic history of these sequences was complex and
marked by duplications and translocations; two instances of transkingdom
horizontal gene transfer were identified. Nonetheless, the sequences fell
within six well-defined evolutionary lineages, three of which contained
only prokaryotes. Of the two prokaryotic/eukaryotic lineages, one contained
bacterial, yeast, slimemold, invertebrate, and vertebrate homologs to human
PGM1 and the second contained likely homologs to human PGM2. Indeed, an
amino acid sequence, derived from a partial human cDNA, that fell within
the second cross-kingdom lineage bears several characteristics expected for
PGM2. A third lineage may contain homologs to human PGM3. On a general
level, our phylogenetic-based approach shows promise for the further
utilization of the extensive molecular database.
相似文献
192.
Oligogalacturonic acids (OGAs), derived from plant cell wall pectin, have
been implicated in a number of signal transduction pathways involved in
growth, development and defense responses of higher plants. This study
investigates the size range of OGAs capable of inducing ethylene synthesis
in tomato plants, and demonstrates that in contrast with many other
effects, only short chain OGAs are active. Oligomers across a range of DP
from 2-15 were separated and purified to homogeneity by QAE-Sephadex anion
exchange chromatography using a novel elution system. The OGAs were applied
to tomato plants and assayed for their ability to induce ethylene gas
release and changes in steady state levels of mRNA encoding the ethylene
forming enzyme aminocyclopropane-1-carboxylic acid oxidase (ACO). The study
demonstrated that only OGAs in the size range of DP4-6 were active both in
eliciting ACO expression and in the production of ethylene.
相似文献
193.
HELENA C. BARROS SOLANGE DA SILVA NEWTON V. VERBISCK MARCIA F. ARAGUTH ROBERTO C. TEDESCO DANIELA O. PROCOPIO RENATO A. MORTARA 《The Journal of eukaryotic microbiology》1996,43(4):275-285
ABSTRACT. Upon incubation at 37° C onto glass coverslips coated with Concanavalin A, poly-L-lysine, or a monoclonal antibody (1D9) directed to the parasite major surface glycoprotein Ssp-4, extracellular Trypanosoma cruzi amastigotes release trails of material barely visible by light microscopy. This release is not associated with parasite movements. Immunolabeling studies confirmed that the material is derived from the parasite's membrane since thin section through samples labeled with 1D9 revealed that the trails are membrane-bound structures. Scanning electron microscopy showed that the ∼0.1-μm thick trails of material emerging from the amastigotes can be uniform or beaded, indicating a tendency to vesiculation. The trails are preferentially released from the flagellar pocket region and/or at the opposite posterior end of the parasite body, and seem to be devoid of microtubules. The release is time and temperature-dependent and fixed parasites do not form trails. All attempts to inhibit trail release using drugs (antimycin A, sodium azide, cytochalasin D, nocodazole, genistein, staurosporine, EGTA) failed. The observation of trails associated with intracellular parasites and amastigotes invading Vero cells suggests that this is probably a physiological process. 相似文献
194.
P Su H Rennert R M Shayiq R Yamamoto Y M Zheng S Addya J F Strauss N G Avadhani 《DNA and cell biology》1990,9(9):657-667
A cDNA expression library prepared from rat liver RNA was screened with a polyclonal antibody specific for mitochondrial vitamin D3 25-hydroxylase and a cDNA for rabbit liver mitochondrial cytochrome P450c26 (CYP 26), yielding cDNA clones with identical sequences. The deduced amino acid sequence derived from a 1.9-kb full-length cDNA was 73% identical to that of rabbit cytochrome P450c26. A monoclonal antibody was used to demonstrate that the product of the 1.9-kb cDNA clone was targeted to the mitochondrial compartment when expressed in COS cells. Mitochondrial membranes containing the expressed protein showed both vitamin D3 25-hydroxylase and cholesterol 26-hydroxylase activities when reconstituted with ferredoxin reductase and ferredoxin, demonstrating that the same P450, designated as P450c26/25, can catalyze both reactions. Northern blot analysis revealed that the P450c26/25 cDNA hybridizes with a 2.4-kb RNA from rat liver and unstimulated ovaries. Treatment of rats with pregnant mare's serum gonadotropin resulted in a fivefold increase in the 2.4-kb mRNA as well as the appearance of a 2.1-kb mRNA species in the ovaries. Our findings document the presence of a regulated bifunctional mitochondrial cytochrome P450 capable of catalyzing the 25-hydroxylation of vitamin D3 and the 26-hydroxylation of cholesterol. 相似文献
195.
JW Mills ADC MacKnight JA Jarrell JM Dayer DA Ausiello 《The Journal of cell biology》1981,88(3):637-643
To determine the specificity and efficacy of [(3)H]ouabain binding as a quantitative measure of the Na(+) pump (Na(+), K(+)-ATPase) and as a marker for the localization of pumps involved in transepithelial Na(+)-transport, we analyzed the interaction of [(3)H]ouabain with its receptor in pig kidney epithelial (LLC-PK(1)) cells. When these epithelial cells are depleted of Na(+) and exposed to 2 muM [(3)H]ouabain in a Na(+)-free medium, binding is reduced by 90 percent. When depleted of K(+) and incubated in a K(+)- free medium, the ouabain binding rate is increase compared with that measured at 5 mM. This increase is only demonstable when Na(+) is present. The increased rate could be attributed to the predominance of the Na(+)-stimulated phosphorylated form of the pump, as K(+) is not readily available to stimulate dephosphorylation. However, some binding in the K(+)-free medium is attributable to pump turnover (and therefore, recycling of K(+)), because analysis of K(+)-washout kinetics demonstrated that addition of 2 muM ouabain to K(+)-depleted cells increased the rate of K(+) loss. These results indicate that in intact epithelial cells, unlike isolated membrane preparations, the most favorable condition for supporting ouabain binding occurs when the Na(+), K(+)-ATPase is operating in the Na(+)-pump mode or is phosphorylated in the presence of Na(+). When LLC-PK(1) cells were exposed to ouabain at 4 degrees C, binding was reduced by 97 percent. Upon rewarming, the rate of binding was greater than that obtained on cells kept at a constant 37 degrees C. However, even at this accelerated rate, the time to reach equilibrium was beyond what is required for cells, swollen by exposure to cold, to recover normal volume. Thus, results from studies that have attempted to use ouabain to eliminate the contribution of the conventional Na(+) pump to volume recovery must be reevaluated if the exposure to ouabain was done in the cold or under conditions in which the Na(+) pump is not operating. 相似文献
196.
MÁRCIA REGINA DA SILVA PEDRINT JOSÉ LUIZ CALDAS WOLFF STEVE REID 《The Annals of applied biology》2004,145(1):107-112
Baculoviruses are a group of viruses that infect invertebrates and that have been used worldwide as a biopesticide against several insect pests of the Order Lepidoptera. In Brazil, the baculovirus Spodoptera frugiperda multicapsid nucleopolyhedrovirus (SfMNPV; Baculoviridae) has been used experimentally to control S. frugiperda (Lepidoptera: Noctuidae), an important insect pest of corn (maize) fields and other crops. Baculoviruses can be produced either in insect larvae or in cell culture bioreactors. A major limitation to the in vitro production of baculoviruses is the rapid generation of mutants when the virus undergoes passages in cell culture. In order to evaluate the potential of in vitro methods of producing SfMNPV on a large-scale, we have multiplied a Brazilian isolate of this virus in cell culture. Extensive formation of few polyhedra mutants was observed after only two passages in Sf9 cells. 相似文献
197.
Adult trees of Quercus petraea were submitted to controlled water shortage in a natural stand near Nancy, France. Diurnal course of net CO2 assimilation rate (A) was measured in situ together with chlorophyll a fluorescence determined on dark adapted leaves. In 1990, trees experienced a strong water stress, with predawn and midday leaf water potentials below –2·0 and –3·0 MPa, respectively. Diurnal course of A of well-watered trees exhibited sometimes important midday decreases in A related to high temperature and vapour pressure deficit. Decreases in initial (Fo) and maximal (Fm) fluorescence and sometimes in photochemical efficiency of photosystem II (Fv/Fm) were observed and probably revealed the onset of mechanisms for thermal de-excitation. These mechanisms were shown to be sensitive to dithiothreitol. All these effects were reversible and vanished almost completely overnight. Therefore, they may be considered as protective mechanisms adjusting activity of photosystem II to the electron requirement for photosynthesis. Water stress amplified these reactions: A was strongly decreased, showing important midday depression; diurnal reductions in Fm and Fv/Fm were enhanced. The same trends were observed during summer 1991, despite a less marked drought. These protective mechanisms seemed very effective, as no photoinhibitory damage to PS II could be detected in either water stressed or control trees. 相似文献
198.
Mutations in the principal neutralization determinant of human immunodeficiency virus type 1 affect syncytium formation, virus infectivity, growth kinetics, and neutralization. 总被引:19,自引:15,他引:4 下载免费PDF全文
R J Grimaila B A Fuller P D Rennert M B Nelson M L Hammarskjld B Potts M Murray S D Putney G Gray 《Journal of virology》1992,66(4):1875-1883
The principal neutralization determinant (PND) of human immunodeficiency virus type 1 envelope glycoprotein gp120 contains a conserved GPG sequence. The effects of a 29-amino-acid deletion of most of the PND, a 3-amino-acid deletion in the GPG sequence, and 16 single-amino-acid substitutions in the GPG sequence were determined in a transient expression assay. All mutant envelope glycoproteins were expressed at levels comparable to that of the wild-type envelope, and mutations in the GPG sequence did not affect processing to gp120 or, except for the 29-amino-acid deletion, binding to CD4. Of all of the mutants, only the GHG and GFG mutants induced formation of syncytia similar in size and number to those induced by the wild-type envelope. When the envelope expression level was increased 10-fold or more, several additional mutants (APG, GAG, GSG, GQG, GVG, and GPF) also induced syncytium formation. Transfection with infectious proviral molecular clones containing the GHG, GFG, APG, GAG, GSG, or GPF mutations induced production of viral particles; however, only the GPG, GHG, and GFG viruses produced active infections in CD4-bearing cells. Furthermore, whereas the wild-type virus was efficiently neutralized by PND polyclonal and monoclonal antibodies, the GHG- and GFG-containing viruses were not. These results show that mutations in the GPG sequence found within the PND do not affect envelope expression and do not significantly affect CD4 binding or production of viral particles but that they do affect the ability of the envelope to induce syncytia and those of the viral particles to infect CD4 cells and be neutralized by PND antibodies. 相似文献
199.
Hydrogen and oxygen isotopic fractionation relative to mediumwater for two different carbohydrate metabolic pathways leadingto cellulose synthesis were measured. This was accomplishedby analysing stable hydrogen and oxygen isotope ratios of waterand cellulose for seedlings. The seedlings had been germinatedand heterotrophically grown in closed vessels from species havingstarch (Triticum aestivum L. and Hordeum vulgare L.) and lipids(Ricinus communisL. and Arachis hypogaea L.) as the primarysubstrate. Isotopic fractionation factors occurring during enzyme-mediatedexchange of carbon-bound hydrogen with water or the additionof carbon-bound hydrogens from water during the synthesis ofcellulose from either starch or lipids were similar (rangingfrom +144 to +166%). About 34% and 67% of carbon-bound hydrogenswere derived from water during the synthesis of cellulose fromstarch and lipid, respectively. Thus, the greater deuteriumenrichment in cellulose from oil seed species associated withgluconeogenesis was caused by a greater proportion of water-derivedcarbon-bound hydrogens and not because of differences in fractionationfactors. The proportion of carbon-bound hydrogens derived fromwater during these metabolic pathways was similar to that ofoxygen derived from water. These results may explain the variabilityin D/H ratios of cellulose nitrate from terrestrial and aquaticplants. Key words: 相似文献
200.
Antiprotease activity in tears and nasal secretions 总被引:1,自引:0,他引:1