全文获取类型
收费全文 | 5005篇 |
免费 | 716篇 |
国内免费 | 1篇 |
出版年
2021年 | 69篇 |
2020年 | 43篇 |
2019年 | 55篇 |
2018年 | 64篇 |
2017年 | 50篇 |
2016年 | 99篇 |
2015年 | 163篇 |
2014年 | 187篇 |
2013年 | 208篇 |
2012年 | 264篇 |
2011年 | 273篇 |
2010年 | 196篇 |
2009年 | 157篇 |
2008年 | 220篇 |
2007年 | 214篇 |
2006年 | 195篇 |
2005年 | 195篇 |
2004年 | 184篇 |
2003年 | 164篇 |
2002年 | 183篇 |
2001年 | 132篇 |
2000年 | 120篇 |
1999年 | 104篇 |
1998年 | 60篇 |
1997年 | 51篇 |
1996年 | 56篇 |
1995年 | 49篇 |
1994年 | 56篇 |
1993年 | 49篇 |
1992年 | 98篇 |
1991年 | 99篇 |
1990年 | 80篇 |
1989年 | 87篇 |
1988年 | 101篇 |
1987年 | 83篇 |
1986年 | 76篇 |
1985年 | 61篇 |
1984年 | 65篇 |
1983年 | 63篇 |
1982年 | 51篇 |
1981年 | 69篇 |
1980年 | 54篇 |
1979年 | 74篇 |
1978年 | 55篇 |
1977年 | 49篇 |
1976年 | 47篇 |
1975年 | 43篇 |
1974年 | 68篇 |
1973年 | 63篇 |
1967年 | 43篇 |
排序方式: 共有5722条查询结果,搜索用时 265 毫秒
971.
Joost Smolders Mari?lle Thewissen Evelyn Peelen Paul Menheere Jan Willem Cohen Tervaert Jan Damoiseaux Raymond Hupperts 《PloS one》2009,4(8)
Background
In several autoimmune diseases, including multiple sclerosis (MS), a compromised regulatory T cell (Treg) function is believed to be critically involved in the disease process. In vitro, the biologically active metabolite of vitamin D has been shown to promote Treg development. A poor vitamin D status has been linked with MS incidence and MS disease activity. In the present study, we assess a potential in vivo correlation between vitamin D status and Treg function in relapsing remitting MS (RRMS) patients.Methodology/Principal Findings
Serum levels of 25-hydroxyvitamin D (25(OH)D) were measured in 29 RRMS patients. The number of circulating Tregs was assessed by flow-cytometry, and their functionality was tested in vitro in a CFSE-based proliferation suppression assay. Additionally, the intracellular cytokine profile of T helper cells was determined directly ex-vivo by flow-cytometry. Serum levels of 25(OH)D correlated positively with the ability of Tregs to suppress T cell proliferation (R = 0.590, P = 0.002). No correlation between 25(OH)D levels and the number of Tregs was found. The IFN-γ/IL-4 ratio (Th1/Th2-balance) was more directed towards IL-4 in patients with favourable 25(OH)D levels (R = −0.435, P = 0.023).Conclusions/Significance
These results show an association of high 25(OH)D levels with an improved Treg function, and with skewing of the Th1/Th2 balance towards Th2. These findings suggest that vitamin D is an important promoter of T cell regulation in vivo in MS patients. It is tempting to speculate that our results may not only hold for MS, but also for other autoimmune diseases. Future intervention studies will show whether modulation of vitamin D status results in modulation of the T cell response and subsequent amelioration of disease activity. 相似文献972.
973.
Jan Felix Drexler Bernd Kupfer Nadine Petersen Rejane Maria Tommasini Grotto Silvia Maria Corvino Rodrigues Klaus Grywna Marcus Panning Augustina Annan Giovanni Faria Silva Jill Douglas Evelyn S. C Koay Heidi Smuts Eduardo M Netto Peter Simmonds Maria Inês de Moura Campos Pardini W. Kurt Roth Christian Drosten 《PLoS medicine》2009,6(2)
Background
Detection and quantification of hepatitis C virus (HCV) RNA is integral to diagnostic and therapeutic regimens. All molecular assays target the viral 5′-noncoding region (5′-NCR), and all show genotype-dependent variation of sensitivities and viral load results. Non-western HCV genotypes have been under-represented in evaluation studies. An alternative diagnostic target region within the HCV genome could facilitate a new generation of assays.Methods and Findings
In this study we determined by de novo sequencing that the 3′-X-tail element, characterized significantly later than the rest of the genome, is highly conserved across genotypes. To prove its clinical utility as a molecular diagnostic target, a prototype qualitative and quantitative test was developed and evaluated multicentrically on a large and complete panel of 725 clinical plasma samples, covering HCV genotypes 1–6, from four continents (Germany, UK, Brazil, South Africa, Singapore). To our knowledge, this is the most diversified and comprehensive panel of clinical and genotype specimens used in HCV nucleic acid testing (NAT) validation to date. The lower limit of detection (LOD) was 18.4 IU/ml (95% confidence interval, 15.3–24.1 IU/ml), suggesting applicability in donor blood screening. The upper LOD exceeded 10−9 IU/ml, facilitating viral load monitoring within a wide dynamic range. In 598 genotyped samples, quantified by Bayer VERSANT 3.0 branched DNA (bDNA), X-tail-based viral loads were highly concordant with bDNA for all genotypes. Correlation coefficients between bDNA and X-tail NAT, for genotypes 1–6, were: 0.92, 0.85, 0.95, 0.91, 0.95, and 0.96, respectively; X-tail-based viral loads deviated by more than 0.5 log10 from 5′-NCR-based viral loads in only 12% of samples (maximum deviation, 0.85 log10). The successful introduction of X-tail NAT in a Brazilian laboratory confirmed the practical stability and robustness of the X-tail-based protocol. The assay was implemented at low reaction costs (US$8.70 per sample), short turnover times (2.5 h for up to 96 samples), and without technical difficulties.Conclusion
This study indicates a way to fundamentally improve HCV viral load monitoring and infection screening. Our prototype assay can serve as a template for a new generation of viral load assays. Additionally, to our knowledge this study provides the first open protocol to permit industry-grade HCV detection and quantification in resource-limited settings. 相似文献974.
Evelyn Gassert Elita Avota Harry Harms Georg Krohne Erich Gulbins Sibylle Schneider-Schaulies 《PLoS pathogens》2009,5(10)
Silencing of T cell activation and function is a highly efficient strategy of immunosuppression induced by pathogens. By promoting formation of membrane microdomains essential for clustering of receptors and signalling platforms in the plasma membrane, ceramides accumulating as a result of membrane sphingomyelin breakdown are not only essential for assembly of signalling complexes and pathogen entry, but also act as signalling modulators, e. g. by regulating relay of phosphatidyl-inositol-3-kinase (PI3K) signalling. Their role in T lymphocyte functions has not been addressed as yet. We now show that measles virus (MV), which interacts with the surface of T cells and thereby efficiently interferes with stimulated dynamic reorganisation of their actin cytoskeleton, causes ceramide accumulation in human T cells in a neutral (NSM) and acid (ASM) sphingomyelinase–dependent manner. Ceramides induced by MV, but also bacterial sphingomyelinase, efficiently interfered with formation of membrane protrusions and T cell spreading and front/rear polarisation in response to β1 integrin ligation or αCD3/CD28 activation, and this was rescued upon pharmacological or genetic ablation of ASM/NSM activity. Moreover, membrane ceramide accumulation downmodulated chemokine-induced T cell motility on fibronectin. Altogether, these findings highlight an as yet unrecognised concept of pathogens able to cause membrane ceramide accumulation to target essential processes in T cell activation and function by preventing stimulated actin cytoskeletal dynamics. 相似文献
975.
Romain Derelle Tsuyoshi Momose Michael Manuel Corinne Da Silva Patrick Wincker Evelyn Houliston 《RNA (New York, N.Y.)》2010,16(4):696-707
Replacement of mRNA 5′ UTR sequences by short sequences trans-spliced from specialized, noncoding, spliced leader (SL) RNAs is an enigmatic phenomenon, occurring in a set of distantly related animal groups including urochordates, nematodes, flatworms, and hydra, as well as in Euglenozoa and dinoflagellates. Whether SL trans-splicing has a common evolutionary origin and biological function among different organisms remains unclear. We have undertaken a systematic identification of SL exons in cDNA sequence data sets from non-bilaterian metazoan species and their closest unicellular relatives. SL exons were identified in ctenophores and in hydrozoan cnidarians, but not in other cnidarians, placozoans, or sponges, or in animal unicellular relatives. Mapping of SL absence/presence obtained from this and previous studies onto current phylogenetic trees favors an evolutionary scenario involving multiple origins for SLs during eumetazoan evolution rather than loss from a common ancestor. In both ctenophore and hydrozoan species, multiple SL sequences were identified, showing high sequence diversity. Detailed analysis of a large data set generated for the hydrozoan Clytia hemisphaerica revealed trans-splicing of given mRNAs by multiple alternative SLs. No evidence was found for a common identity of trans-spliced mRNAs between different hydrozoans. One feature found specifically to characterize SL-spliced mRNAs in hydrozoans, however, was a marked adenosine enrichment immediately 3′ of the SL acceptor splice site. Our findings of high sequence divergence and apparently indiscriminate use of SLs in hydrozoans, along with recent findings in other taxa, indicate that SL genes have evolved rapidly in parallel in diverse animal groups, with constraint on SL exon sequence evolution being apparently rare. 相似文献
976.
P. D. Cox L. Matthews R. J. Jacobson R. Cannon A. MacLeod K. F. A. Walters 《Biocontrol Science and Technology》2006,16(9):871-891
Thrips palmi is a major pest of many crops in the tropics and sub-tropics, and is a serious threat within the protected horticulture industry in other parts of the world including the UK. Widespread use of insecticides against T. palmi throughout the world coupled with the restricted range of products available makes it essential to find alternative systems for control. The scattered information on its natural enemies, particularly predators and parasitoids, is reviewed and their potential for use in the control of T. palmi as part of IPM strategies in the UK is considered. Natural enemies selected for detailed examination include: Amblyseius spp., Anthocoris nemoralis, Atheta coriaria, Bilia spp., Campylomma spp., Ceranisus spp., Deraeocoris spp., Franklinothrips spp., Hypoaspis spp., Orius spp. and Phytoseius spp. Recommendations for further investigations are made, including screening and efficacy testing of candidate predators and parasitoids, using semiochemicals to enhance their effectiveness, and assessing the compatibility of chosen species with other components of an IPM system. 相似文献
977.
978.
Polyketide synthases cannot be functional unless their apo-acyl carrier proteins (apo-ACPs) are post-translationally modified by covalent attachment of the 4'-phosphopantetheine group to the highly conserved serine residue, and this reaction is catalyzed by phosphopantetheinyl transferases (PPTases). Cloning and sequence analysis of the 33-kb fredericamycin (FDM) biosynthetic gene cluster from Streptomyces griseus revealed fdmW, whose deduced gene product showed significant sequence homology to known PPTases. Biochemical characterization of FdmW in vitro confirmed that it is a PPTase. Inactivation of fdmW resulted in approximately 93% reduction of FDM production, and complementation of the fdmW::aac (3)IV mutant by expressing fdmW in trans restored FDM production to a level comparable with that of the wild-type strain. Although FdmW can phosphopantetheinylate various ACPs, it prefers its cognate substrate, the FdmH ACP, with a K(m) of 5.8 microM and a k(cat)/K(m) of 8.1 microM(-1) x min(-1), to heterologous ACPs, such as the TcmM ACP with a K(m) of 1.0 x 10(2) microM and a k(cat) /K(m) of 0.6 microM(-1) x min(-1). These findings suggest that FdmW is specific for FDM biosynthesis. FdmW therefore represents the first holo-ACP synthase-type PPTase identified from an aromatic polyketide biosynthetic gene cluster. 相似文献
979.
Fox KL Cox AD Gilbert M Wakarchuk WW Li J Makepeace K Richards JC Moxon ER Hood DW 《The Journal of biological chemistry》2006,281(52):40024-40032
The lipopolysaccharide (LPS) of non-typeable Haemophilus influenzae (NTHi) can be substituted at various positions by N-acetylneuraminic acid (Neu5Ac). LPS sialylation plays an important role in pathogenesis. The only LPS sialyltransferase characterized biochemically to date in H. influenzae is Lic3A, an alpha-2,3-sialyltransferase responsible for the addition of Neu5Ac to a lactose acceptor (Hood, D. W., Cox, A. D., Gilbert, M., Makepeace, K., Walsh, S., Deadman, M. E., Cody, A., Martin, A., M?nsson, M., Schweda, E. K., Brisson, J. R., Richards, J. C., Moxon, E. R., and Wakarchuk, W. W. (2001) Mol. Microbiol. 39, 341-350). Here we describe a second sialyltransferase, Lic3B, that is a close homologue of Lic3A and present in 60% of NTHi isolates tested. A recombinant form of Lic3B was expressed in Escherichia coli and purified by affinity chromatography. We used synthetic fluorescent acceptors with a terminal lactose or sialyllactose to show that Lic3B has both alpha-2,3- and alpha-2,8-sialyltransferase activities. Structural analysis of LPS from lic3B mutant strains of NTHi confirmed that only monosialylated species were detectable, whereas disialylated species were detected upon inactivation of lic3A. Furthermore, introduction of lic3B into a lic3B-deficient strain background resulted in a significant increase in sialylation in the recipient strain. Mass spectrometric analysis of LPS indicated that glycoforms containing two Neu5Ac residues were evident that were not present in the LPS of the parent strain. These findings characterize the activity of a second sialyltransferase in H. influenzae, responsible for the addition of di-sialic acid to the LPS. Modification of the LPS by di-sialylation conferred increased resistance of the organism to the killing effects of normal human serum, as compared with mono-sialylated or non-sialylated species, indicating that this modification has biological significance. 相似文献
980.
Walter Jaoko Etienne Karita Kayitesi Kayitenkore Gloria Omosa-Manyonyi Susan Allen Soe Than Elizabeth M. Adams Barney S. Graham Richard A. Koup Robert T. Bailer Carol Smith Len Dally Bashir Farah Omu Anzala Claude M. Muvunyi Jean Bizimana Tony Tarragona-Fiol Philip J. Bergin Peter Hayes Martin Ho Kelley Loughran Wendy Komaroff Gwynneth Stevens Helen Thomson Mark J. Boaz Josephine H. Cox Claudia Schmidt Jill Gilmour Gary J. Nabel Patricia Fast Job Bwayo 《PloS one》2010,5(9)