全文获取类型
收费全文 | 15719篇 |
免费 | 1395篇 |
国内免费 | 1589篇 |
出版年
2024年 | 27篇 |
2023年 | 193篇 |
2022年 | 518篇 |
2021年 | 862篇 |
2020年 | 652篇 |
2019年 | 746篇 |
2018年 | 712篇 |
2017年 | 527篇 |
2016年 | 671篇 |
2015年 | 995篇 |
2014年 | 1153篇 |
2013年 | 1278篇 |
2012年 | 1534篇 |
2011年 | 1287篇 |
2010年 | 792篇 |
2009年 | 759篇 |
2008年 | 826篇 |
2007年 | 692篇 |
2006年 | 609篇 |
2005年 | 556篇 |
2004年 | 399篇 |
2003年 | 397篇 |
2002年 | 358篇 |
2001年 | 245篇 |
2000年 | 208篇 |
1999年 | 240篇 |
1998年 | 134篇 |
1997年 | 147篇 |
1996年 | 137篇 |
1995年 | 122篇 |
1994年 | 117篇 |
1993年 | 91篇 |
1992年 | 102篇 |
1991年 | 75篇 |
1990年 | 84篇 |
1989年 | 63篇 |
1988年 | 60篇 |
1987年 | 44篇 |
1986年 | 50篇 |
1985年 | 53篇 |
1984年 | 35篇 |
1983年 | 23篇 |
1982年 | 23篇 |
1981年 | 11篇 |
1980年 | 10篇 |
1979年 | 16篇 |
1978年 | 6篇 |
1977年 | 12篇 |
1976年 | 9篇 |
1973年 | 9篇 |
排序方式: 共有10000条查询结果,搜索用时 156 毫秒
971.
972.
Karimi-Busheri F Marcoux Y Tredget EE Li L Zheng J Ghoreishi M Weinfeld M Ghahary A 《Journal of cellular biochemistry》2002,86(4):737-747
Annexin II is a multifunctional calcium-dependent phospholipid binding protein whose presence in epidermis has previously been reported. However, like other members of annexin family, annexin II has been regarded as either an intracellular protein or associated with the cellular membrane. Here, we report the presence of a releasable annexin II and p11, two monomers of annexin II tetramer, in keratinocyte-conditioned medium (KCM). Proteins present in KCM were fractionated on a gel filtration column and following further evaluation, a releasable protein with apparent MW of 36 kDa was identified. Further characterization identified this protein as the p36 monomer of annexin II tetramer. The phospho-tyrosine antibody did not visualize this protein as the phosphorylated form of p36. Several experiments were conducted to examine whether this protein is soluble or associated with keratinocyte cell membranes in the conditioned medium. A centrifugation of conditioned medium was not able to bring this protein down into the pellet. Surprisingly, the results of Western analysis identified p36 and p11, two monomers of the annexin II tetramer, in conditioned medium derived from either keratinocytes cultured alone or keratinocytes co-cultured with fibroblasts. In contrast to the keratinocyte-conditioned medium in which annexin II was easily detectable, both monomers were barely detectable in conditioned medium collected from dermal fibroblasts. This finding was in contrast to the cell lysates in which p36 was detectable in both keratinocytes and fibroblasts. However, the amount of this protein was markedly higher in keratinocyte lysate relative to that of dermal fibroblasts. Conditioned medium derived from keratinocyte established from adult showed a higher level of annexin II compared to that of keratinocytes established from newborn babies. The expression of p11 seems to increase with differentiation of keratinocytes derived from either adult or newborn skin samples. When the site of annexin synthesis in human skin was examined by immunohistochemical staining, the antibody for p36 localized the annexin to the keratinocyte cell members in the basal and suprabasal keratinocytes. In conclusion, Western blot detection of both p36 and p11 in conditioned medium from skin cells revealed that human keratinocytes, but not fibroblasts, express a releasable monomer form of annexin II which is regulated by differentiation status of keratinocytes. This finding is consistent with the localization of annexin II detected by immunohistochemical staining. 相似文献
973.
A decade of differential display 总被引:23,自引:0,他引:23
Liang P 《BioTechniques》2002,33(2):338-44, 346
It has been 10 years since the invention of differential display (DD), a conceptually simple methodology that allows the detection and identification of differentially expressed genes. In the past decade, the number of publications describing successful applications of DD has outnumbered those using any other competing methodologies, including subtractive hybridization, representational difference analysis, serial analysis of gene expression, and DNA microarrays. This review will provide a glimpse of the current progress made in DD technological development, refinement, and automation. Excellent examples of DD applications in studying a variety of biological problems, in such diverse biological systems as bacteria, yeast, flies, plants, and higher mammals, are presented to provide a roadmap for those who would like to pursue a fruitful gene "fishing" expedition. Some of the fundamental differences between DD and DNA microarrays are also discussed. 相似文献
974.
In chromosomes of metazoa, the assembly of the genome into chromatin makes an important but poorly understood contribution to determining where DNA replication will initiate. We addressed this issue by studying the developmental progression of the location of the DNA replication origin (ORI) and alterations in chromatin structure in one of the best-mapped ORIs in metazoa, that found in DNA puff II/9A of the fly Sciara coprophila. We found that DNA synthesis for both normal chromosomal endoduplication and DNA amplification initiates within the same 5.5 kb EcoRI fragment. We showed that irrespective of the mode of ORI function--replication or amplification--chromatin over the 1 kb major ORI is never remodeled into a conventional DNase I hypersensitive site (DH site). Instead, we found that the major site of alterations to chromatin structure at this locus is a large (approximately 400 bp) DH site located 600 bp away from the major ORI, at a position where the frequency of replication initiation events falls dramatically. We describe a tight positive correlation between ORI activity, strength of this DH site, and the intranuclear titer of protein factor(s) that bind the DH site in a sequence-specific manner. We propose that the Sciara replicator in locus II/9A is composed of sequences that reside within the ORI per se as well as sequences encompassed by the DH site. 相似文献
975.
Andrew V. Suarez David A. HolwayDangsheng Liang Neil D. Tsutsui Ted J. Case 《Animal behaviour》2002,64(5):697-708
The Argentine ant, Linepithema humile, is a widespread invasive species characterized by reduced intraspecific aggression within introduced populations. To illuminate the mechanisms underlying nestmate recognition in Argentine ants, we studied the spatial and temporal fidelity of intraspecific aggression in an introduced population of Argentine ants within which intraspecific aggression does occur. We quantified variation in the presence or absence of intraspecific aggression among nests over time both in the field and under controlled laboratory conditions to gain insight into the role of environmental factors as determinants of nestmate discriminatory ability. In addition, we compared levels of intraspecific aggression between nest pairs to the similarity of their cuticular hydrocarbons to determine the potential role of these compounds as labels for nestmate discrimination. In both field and laboratory comparisons, nest pairs behaved in a consistent manner throughout the course of the experiment: pairs that fought did so for an entire year, and pairs that did not fight remained nonaggressive. Moreover, we found a negative relationship between cuticular hydrocarbon similarity and the degree of aggression between nests, suggesting that these hydrocarbons play a role in nestmate discriminatory ability. In contrast to the prevailing pattern, ants from one site showed a marked change in behaviour during the course of this study. A concomitant change was also seen in the cuticular hydrocarbon profiles of ants from this site. Copyright 2002 The Association for the Study of Animal Behaviour. Published by Elsevier Science Ltd. All rights reserved. 相似文献
976.
GAP-43 (neuromodulin) is a protein kinase C substrate that is abundant in developing and regenerating neurons. Thioester-linked palmitoylation at two cysteines near the GAP-43 N terminus has been implicated in directing membrane binding. Here, we use mass spectrometry to examine the stoichiometry of palmitoylation and the molecular identity of the fatty acid(s) attached to GAP-43 in vivo. GAP-43 expressed in either PC12 or COS-1 cells was acetylated at the N-terminal methionine. Approximately 35% of the N-terminal GAP-43 peptides were also modified by palmitate and/or stearate on Cys residues. Interestingly, a variety of acylated species was detected, in which one of the Cys residues was acylated by either palmitate or stearate, or both Cys residues were acylated by palmitates or stearates or a combination of palmitate and stearate. Depalmitoylation of membrane-bound GAP-43 did not release the protein from the membrane, implying that additional forces function to maintain membrane binding. Indeed, mutation of four basic residues within the N-terminal domain of GAP-43 dramatically reduced membrane localization of GAP-43 without affecting palmitoylation. These data reveal the heterogeneous nature of S-acylation in vivo and illustrate the power of mass spectrometry for identification of key regulatory protein modifications. 相似文献
977.
978.
Van Hooser JP Liang Y Maeda T Kuksa V Jang GF He YG Rieke F Fong HK Detwiler PB Palczewski K 《The Journal of biological chemistry》2002,277(21):19173-19182
The visual process is initiated by the photoisomerization of 11-cis-retinal to all-trans-retinal. For sustained vision the 11-cis-chromophore must be regenerated from all-trans-retinal. This requires RPE65, a dominant retinal pigment epithelium protein. Disruption of the RPE65 gene results in massive accumulation of all-trans-retinyl esters in the retinal pigment epithelium, lack of 11-cis-retinal and therefore rhodopsin, and ultimately blindness. We reported previously (Van Hooser, J. P., Aleman, T. S., He, Y. G., Cideciyan, A. V., Kuksa, V., Pittler, S. J., Stone, E. M., Jacobson, S. G., and Palczewski, K. (2000) Proc. Natl. Acad. Sci. U. S. A. 97, 8623-8628) that in Rpe65-/- mice, oral administration of 9-cis-retinal generated isorhodopsin, a rod photopigment, and restored light sensitivity to the electroretinogram. Here, we provide evidence that early intervention by 9-cis-retinal administration significantly attenuated retinal ester accumulation and supported rod retinal function for more than 6 months post-treatment. In single cell recordings rod light sensitivity was shown to be a function of the amount of regenerated isorhodopsin; high doses restored rod responses with normal sensitivity and kinetics. Highly attenuated residual rod function was observed in untreated Rpe65-/- mice. This rod function is likely a consequence of low efficiency production of 11-cis-retinal by photo-conversion of all-trans-retinal in the retina as demonstrated by retinoid analysis. These studies show that pharmacological intervention produces long lasting preservation of visual function in dark-reared Rpe65-/- mice and may be a useful therapeutic strategy in recovering vision in humans diagnosed with Leber congenital amaurosis caused by mutations in the RPE65 gene, an inherited group of early onset blinding and retinal degenerations. 相似文献
979.
Yoshie Matsuda Genqing Liang Yali Zhu Fengshan Ma Richard S. Nelson Biao Ding 《Protoplasma》2002,220(1-2):0051-0058
Summary. Previous work has demonstrated that some endogenous plant gene promoters are active in selective companion cells of the phloem,
depending on organ types and developmental stages. Here we report that the Commelina yellow mottle virus (CoYMV) promoter
is active in the companion cells of leaves, stems and roots of transgenic Nicotiana tabacum cv. Xanthi NN, using β-glucuronidase (GUS) as a reporter. Thus, the CoYMV promoter has a broad organ specificity. This promoter
can be useful in molecular studies on the functions of companion cells in many aspects of phloem biology, such as regulation
of long-distance transport, macromolecular traffic, plant development and interaction with pathogens. It may also be useful
in engineering crops that produce specific gene products in the companion cells to block long-distance movement of pathogens.
Received February 5, 2002; accepted March 27, 2002; published online July 4, 2002
RID="*"
ID="*" Correspondence and reprints: Department of Plant Biology and Plant Biotechnology Center, 207 Rightmire Hall, Ohio State
University, 1060 Carmack Road, Columbus, OH 43210, U.S.A. 相似文献
980.
Calcium removal from the cytoplasm was investigated in freshly isolated aortic endothelial cells by monitoring changes in intracellular calcium ([Ca(2+)](i)) using ratiometric fura-2 fluorimetry. Blockade of the Na(+)/Ca(2+) exchanger (NCX) by replacement of external sodium with equi-molar N-methyl-D-glutamine (0Na PSS) decreased the removal rate by 52%. Blockade of the sarco/endoplasmic reticulum Ca(2+) ATPase (SERCA) by cyclopiazonic acid (CPA) decreased the removal rate by 50%. Simultaneous application of CPA and 0Na PSS did not reduce the removal rate any further (53%). The lack of additivity of these two procedures, suggests that SERCA and the NCX function in series to lower [Ca(2+)](i). In addition, in the absence of extracellular Ca(2+), removal of external Na(+) markedly reduced the rate of loss of Ca(2+) from the ER further supporting the hypothesis that NCX is functionally linked to ER calcium release channels, and thus, plays an important role in ER calcium unloading. To investigate the mechanism for the coupling of NCX and SERCA, the same protocols as described above were repeated after treating the cells with cytochalasin D, which disrupts the cytoskeleton. This treatment uncoupled the NCX from SERCA, as evidenced by the resulting additive inhibitory effects of application of CPA and removal of extracellular Na(+) on the rate of Ca(2+) removal from the cytoplasm. These data suggest that in endothelial cells NCX and SERCA function in series to remove about half of the free Ca(2+) from the cytosol, while PMCA contributes to the other half of the Ca(2+) removal process. 相似文献