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811.
Dr. Juliet Morgan Louis Cohen 《In vitro cellular & developmental biology. Plant》1974,10(3-4):188-195
Summary This report describes in detail a method of enzymatic separation of adult mammalian muscle using papain. The procedure has
proved valuable in the preparation of suspensions of muscle cell pieces from normal human skeletal muscle obtained from patients
of all ages, from 3 months to 79 years. Muscle cultures have been successfully growth from biopsy material from boys with
Duchenne’s muscular dystrophy and from their mothers. The procedure was initially established with adult canine skeletal muscle
and has also been used for monkey muscle.
Small pieces of skeletal muscle are chopped in a solution of 0.05% papain and 0.01% cysteine hydrochloride in Ca2+-and Mg2+-free balanced salt solution and transferred in the papain solution to a flask, in which they are incubated at 37°C for 10
min with occasional agitation. The resulting cell suspension is collected and the remaining pieces are treated with further
portions of fresh papain until only connective tissue remains. The cell pellets obtained by centrifugation are resuspended
in Eagle’s minimum essential medium (supplemented with 20% fetal calf serum) and transferred to culture chambers. The muscle
can be observed at all times, during the separation procedure and subsequently in culture. The events occurring during skeletal
muscle regeneration can be followed. Using the same papain preparation, myoblasts and myotubes may be subcultured and collected
for indefinite frozen storage in dimethylsulfoxide.
This work was supported by a grant-in-aid from the American Heart Association with funds contributed in part by the Chicago
Heart Association, the Pharmaceutical Manufactures Association, and National Institutes of Health Research Grant NS 10385
from the Institute of Neurological Diseases and Stroke. 相似文献
812.
Louis Milakofsky Theodore A. Hare Joanne M. Miller Wolfgang H. Vogel 《Life sciences》1984,34(14):1333-1340
A novel, sensitive and highly resolving amino acid analysis procedure was developed and used to compare two methods of obtaining blood from experimental animals. The procedure, utilizing a meter long microbore HPLC column containing spherical cation-exchange resin and fluorescence detection following postcolumn reaction with o-phthaldialdehyde, was shown to reliably measure forty-one primary amine components in rat plasma.Comparison of values from blood obtained by decapitation and by catheterization documented the significant artifactual influence of the decapitation procedure on approximately half of the measured constituents. 相似文献
813.
814.
The role of Halobacterium cutirubrum deoxyribonucleic acid-dependent ribonucleic acid polymerase subunits in initiation and polymerization 总被引:5,自引:3,他引:2 下载免费PDF全文
1. The two subunits alpha and beta of Halobacterium cutirubrum DNA-dependent RNA polymerase are required in equimolar amounts for RNA synthesis to occur in vitro at the maximum rate. 2. In the absence of bivalent cations no interaction occurs between alpha and beta subunits or between the subunits and DNA. 3. Mn(2+) causes the subunits to form a 1:1 complex that still does not bind to the template. 4. Mg(2+) permits binding of the Mn(2+)-mediated complex to DNA. 5. The complete enzyme, alphabeta, is inhibited by rifampicin and only the beta subunit relieves the inhibition when added in excess. 6. Rifampicin-insensitive, template-dependent RNA synthesis occurs in the presence of protein alpha alone provided an oligonucleotide with a 5'-purine terminus is supplied as primer. 7. In the primed reaction with the alpha protein and an oligonucleotide, the template specificity is independent of the ionic strength, in contrast with the marked effect of salt concentration on the template specificity of the complete enzyme. 8. It is concluded that the beta protein controls the specificity of chain initiation and the template specificity of the complete enzyme and also carries the rifampicin-binding site, whereas the catalytic site is on the alpha subunit. 相似文献
815.
816.
Considerable research efforts have focused on elucidating the systematic relationships among salmonid fishes; an understanding of these patterns of relatedness will inform conservation- and fisheries-related issues, as well as provide a framework for investigating evolutionary mechanisms in the group. However, uncertainties persist in current Salmonidae phylogenies due to biological and methodological factors, and a comprehensive phylogeny including most representatives of the family could provide insight into the causes of these difficulties. Here we increase taxon sampling by including nearly all described salmonid species (n = 63) to present a time-calibrated and more complete portrait of Salmonidae using a combination of molecular markers and analytical techniques. This strategy improved resolution by increasing the signal-to-noise ratio and helped discriminate methodological and systematic errors from sources of difficulty associated with biological processes. Our results highlight novel aspects of salmonid evolution. First, we call into question the widely-accepted evolutionary relationships among sub-families and suggest that Thymallinae, rather than Coregoninae, is the sister group to the remainder of Salmonidae. Second, we find that some groups in Salmonidae are older than previously thought and that the mitochondrial rate of molecular divergence varies markedly among genes and clades. We estimate the age of the family to be 59.1 MY (CI: 63.2-58.1 MY) old, which likely corresponds to the timing of whole genome duplication in salmonids. The average, albeit highly variable, mitochondrial rate of molecular divergence was estimated as ∼0.31%/MY (CI: 0.27–0.36%/MY). Finally, we suggest that some species require taxonomic revision, including two monotypic genera, Stenodus and Salvethymus. In addition, we resolve some relationships that have been notoriously difficult to discern and present a clearer picture of the evolution of the group. Our findings represent an important contribution to the systematics of Salmonidae, and provide a useful tool for addressing questions related to fundamental and applied evolutionary issues. 相似文献
817.
Short-timescale variability of picophytoplankton abundance and cellular parameters in surface waters of the Alboran Sea (western Mediterranean) 总被引:1,自引:0,他引:1
Jacquet Stephan; Prieur Louis; Avois-Jacquet Carol; Lennon Jean-Francois; Vaulot Daniel 《Journal of plankton research》2002,24(7):635-651
The Alboran Sea (western Mediterranean) is characterized bya well-defined hydrological structure, the Almeria (Spain)Oran(Algeria) geostrophic front. During the Almofront-2 cruise (November22, 1997 to January 18, 1998), high frequency sampling ( 相似文献
818.
The Dicer1 allele is used to show that microRNAs (miRNAs) play important roles in astrocyte development and functions. While it is known that astrocytes that lack miRNAs are dysregulated, the in vivo phenotypes of these astrocytes are not well understood. In this study, we use Aldh1l1-EGFP transgene, a marker of astrocytes, to characterize mouse models with conditional Dicer1 ablation (via either human or mouse GFAP-Cre). This transgene revealed novel features of the defective astrocytes from the absence of miRNA. Although astrocyte miRNAs were depleted in both lines, we found histological and molecular differences in the Aldh1l1-EGFP cells between the two Cre lines. Aldh1l1-EGFP cells from hGFAP-Cre mutant lines displayed up-regulation of Aldh1l1-EGFP with increased proliferation and a genomic profile that acquired many features of wildtype primary astrocyte cultures. In the young mGFAP-Cre mutant lines we found that Aldh1l1-EGFP cells were disorganized and hyperproliferative in the developing cerebellum. Using the Aldh1l1-EGFP transgene, our work provides new insights into the roles of miRNAs in astrocyte development and the features of astrocytes in these two mouse models. 相似文献
819.
Polystoma claudecombesi is described as a new species of the Polystomatidae (Monogenea) parasitic in the urinary bladder of the anuran host Rana angolensis. This parasite was collected at three localities in South Africa, namely Vernon Crookes Nature Reserve in Natal province,
Witsieshoek Mountain Resort in the north-eastern Orange Free State and Bovenste Oog in the province of Transvaal. It is the
largest African polystome described to date. Prevalence at Vernon Crookes Nature Reserve was 4.0% and 2.7% in two successive
years, with a mean intensity of 1.0 (n=25 and 36, respectively). One of the two R. angolensis from Witsieshoek was infected with 5 parasites. The diversity and distribution of southern African polystomes is also discussed. 相似文献
820.
Ylenia Chiari Pablo Orozco-terWengel Miguel Vences David R. Vieites Augustin Sarovy Jasmin E. Randrianirina Axel Meyer Edward Louis Jr. 《Conservation Genetics》2006,7(4):473-482
Dyscophus antongilii and D. guineti are two morphologically very similar microhylid frogs from Madagascar of uncertain taxonomy. D. antongilii is currently included in Appendix I of the Convention on the International Trade in Endangered Species (CITES) and its exportation is banned completely. In contrast, D. guineti does not receive any legal protection and it is regularly exported. Field data on ecology and behaviour are to a large extent lacking. Here we report on a genetic survey of D. antongilii and D. guineti using nuclear and mitochondrial DNA markers. Sequences of a fragment of 501 bp of the mitochondrial cytochrome b gene from one population of D. antongilii and two populations of D. guineti resulted in a single haplotype network, without haplotype sharing among the populations. However, haplotypes of D.␣guineti were only 1–4 mutational steps from those of D. antongilii, and did not form a clade. The analysis of eight microsatellites newly developed and standardized for D. antongilii revealed an excess of homozygotes and the absence of Hardy–Weinberg equilibrium. The microsatellite data clearly distinguished between D. antongilii and D. guineti, and fixed differences were observed at one locus. Although confirmation of the status of Dyscophus antongilii and D. guineti as separate species requires further data, our study supports the definition of these two taxa as different evolutionary significant units under the adaptive evolutionary conservation concept. 相似文献