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41.
42.
Valentina Ferrari Alison Tarke Hannah Fields Luca Ferrari Trevor Conley Franco Ferrari Zeynep Koşaloğlu-Yalçın Alessandro Sette Bjoern Peters Colin L. McCarthy Asad Bashey Dimitrios Tzachanis Edward D. Ball Tiffany N. Tanaka Rafael Bejar Thomas A. Lane Antonella Vitiello 《Cytotherapy》2021,23(4):320-328
Therapies that utilize immune checkpoint inhibition work by leveraging mutation-derived neoantigens and have shown greater clinical efficacy in tumors with higher mutational burden. Whether tumors with a low mutational burden are susceptible to neoantigen-targeted therapy has not been fully addressed. To examine the feasibility of neoantigen-specific adoptive T-cell therapy, the authors studied the T-cell response against somatic variants in five patients with myelodysplastic syndrome (MDS), a malignancy with a very low tumor mutational burden. DNA and RNA from tumor (CD34+) and normal (CD3+) cells isolated from the patients’ blood were sequenced to predict patient-specific MDS neopeptides. Neopeptides representing the somatic variants were used to induce and expand autologous T cells ex vivo, and these were systematically tested in killing assays to determine the proportion of neopeptides yielding neoantigen-specific T cells. The authors identified a total of 32 somatic variants (four to eight per patient) and found that 21 (66%) induced a peptide-specific T-cell response and 19 (59%) induced a T-cell response capable of killing autologous tumor cells. Of the 32 somatic variants, 11 (34%) induced a CD4+ response and 11 (34%) induced a CD8+ response that killed the tumor. These results indicate that in vitro induction of neoantigen-specific T cells is feasible for tumors with very low mutational burden and that this approach warrants investigation as a therapeutic option for such patients. 相似文献
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44.
Stephen Millam Alan T. H. Burns Trevor J. Hocking 《Plant Cell, Tissue and Organ Culture》1991,24(1):43-47
The effects of three different general purification protocols have been assessed quantitatively using mesophyll protoplasts of Brassica napus. Within the initial sample two distinct sub-populations were determined. The methods used influenced the ratio of the vacuolated to chloroplastic type protoplast sub-populations. Overall recovery rates of the initial sample varied according to the method used from 38% to 27%, but the relative recovery of the sub-populations varied considerably with a purified ratio of between 1.0:0.78 to 1.0:7.0. Size distribution profiles of the initial and purified populations are also presented. 相似文献
45.
Expression of Ca2+-permeable two-pore channels rescues NAADP signalling in TPC-deficient cells 下载免费PDF全文
Margarida Ruas Lianne C Davis Cheng-Chang Chen Anthony J Morgan Kai-Ting Chuang Timothy F Walseth Christian Grimm Clive Garnham Trevor Powell Nick Platt Frances M Platt Martin Biel Christian Wahl-Schott John Parrington Antony Galione 《The EMBO journal》2015,34(13):1743-1758
The second messenger NAADP triggers Ca2+ release from endo-lysosomes. Although two-pore channels (TPCs) have been proposed to be regulated by NAADP, recent studies have challenged this. By generating the first mouse line with demonstrable absence of both Tpcn1 and Tpcn2 expression (Tpcn1/2−/−), we show that the loss of endogenous TPCs abolished NAADP-dependent Ca2+ responses as assessed by single-cell Ca2+ imaging or patch-clamp of single endo-lysosomes. In contrast, currents stimulated by PI(3,5)P2 were only partially dependent on TPCs. In Tpcn1/2−/− cells, NAADP sensitivity was restored by re-expressing wild-type TPCs, but not by mutant versions with impaired Ca2+-permeability, nor by TRPML1. Another mouse line formerly reported as TPC-null likely expresses truncated TPCs, but we now show that these truncated proteins still support NAADP-induced Ca2+ release. High-affinity [32P]NAADP binding still occurs in Tpcn1/2−/− tissue, suggesting that NAADP regulation is conferred by an accessory protein. Altogether, our data establish TPCs as Ca2+-permeable channels indispensable for NAADP signalling. 相似文献
46.
Human type 3 3alpha-hydroxysteroid dehydrogenase, or aldo-keto reductase (AKR) 1C2, eliminates the androgen signal in human prostate by reducing 5alpha-dihydrotestosterone (DHT, potent androgen) to form 3alpha-androstanediol (inactive androgen), thereby depriving the androgen receptor of its ligand. The k(cat) for the NADPH-dependent reduction of DHT catalyzed by AKR1C2 is 0.033 s(-1). We employed transient kinetics and kinetic isotope effects to dissect the contribution of discrete steps to this low k(cat) value. Stopped-flow experiments to measure the formation of the AKR1C2.NADP(H) binary complex indicated that two slow isomerization events occur to yield a tight complex. A small primary deuterium isotope effect on k(cat) (1.5) and a slightly larger effect on k(cat)/K(m) (2.1) were observed in the steady state. In the transient state, the maximum rate constant for the single turnover of DHT (k(trans)) was determined to be 0.11 s(-1) for the NADPH-dependent reaction, which was approximately 4-fold greater than the corresponding k(cat) x k(trans) was significantly reduced when NADPD was substituted for NADPH, resulting in an apparent (D)k(trans) of 3.5. Thus, the effects of isotopic substitution on the hydride transfer step were masked by slow events that follow or precede the chemical transformation. Transient multiple-turnover reactions generated curvilinear reaction traces, consistent with the product formation and release occurring at comparable rates. Global fitting analysis of the transient kinetic data enabled the estimate of the rate constants for the three-step cofactor binding/release model and for the minimal ordered bi-bi turnover mechanism. Results were consistent with a kinetic mechanism in which a series of slow events, including the chemical step (0.12 s(-1)), the release of the steroid product (0.081 s(-1)), and the release of the cofactor product (0.21 s(-1)), combine to yield the overall observed low turnover number. 相似文献
47.
Kim Y. C. Fung Bruce Tabor Michael J. Buckley Ilka K. Priebe Leanne Purins Celine Pompeia Gemma V. Brierley Trevor Lockett Peter Gibbs Jeanne Tie Paul McMurrick James Moore Andrew Ruszkiewicz Edouard Nice Timothy E. Adams Antony Burgess Leah J. Cosgrove 《PloS one》2015,10(3)
Background
The majority of colorectal cancer (CRC) cases are preventable by early detection and removal of precancerous polyps. Even though CRC is the second most common internal cancer in Australia, only 30 per cent of the population considered to have risk factors participate in stool-based test screening programs. Evidence indicates a robust, blood-based, diagnostic assay would increase screening compliance. A number of potential diagnostic blood-based protein biomarkers for CRC have been reported, but all lack sensitivity or specificity for use as a stand-alone diagnostic. The aim of this study was to identify and validate a panel of protein-based biomarkers in independent cohorts that could be translated to a reliable, non-invasive blood-based screening test.Principal Findings
In two independent cohorts (n = 145 and n = 197), we evaluated seven single biomarkers in serum of CRC patients and age/gender matched controls that showed a significant difference between controls and CRC, but individually lack the sensitivity for diagnostic application. Using logistic regression strategies, we identified a panel of three biomarkers that discriminated between controls and CRC with 73% sensitivity at 95% specificity, when applied to either of the two cohorts. This panel comprised of Insulin like growth factor binding protein 2 (IGFBP2), Dickkopf-3 (DKK3), and Pyruvate kinase M2(PKM2).Conclusions
Due to the heterogeneous nature of CRC, a single biomarker is unlikely to have sufficient sensitivity or specificity for use as a stand-alone diagnostic screening test and a panel of markers may be more effective. We have identified a 3 biomarker panel that has higher sensitivity and specificity for early stage (Stage I and -II) disease than the faecal occult blood test, raising the possibility for its use as a non-invasive blood diagnostic or screening test. 相似文献48.
The mitochondrion is an essential cellular compartment in eukaryotes. The mitochondrial proteins Tom20 and Tom22 are receptors that ensure recognition and binding of proteins imported for mitochondrial biogenesis. Comparison of the sequence for the Tom20 and Tom22 subunits in the yeasts Saccharomyces cerevisiae and Saccharomyces castellii, show a rare case of domain stealing, where in Saccharomyces castellii Tom22 has lost an acidic domain, and Tom20 has gained one. This example of domain stealing is a snapshot of evolution in action and provides excellent evidence that Tom20 and Tom22 are subunits of a single, composite receptor that binds precursor proteins for import into mitochondria. 相似文献
49.
Schallmey M Ly A Wang C Meglei G Voget S Streit WR Driscoll BT Charles TC 《FEMS microbiology letters》2011,322(2):150-156
Salmonella enterica serovar Typhi and Typhimurium are closely related serovars. However, S. Typhi, a human-specific pathogen, has 5% of genes as pseudogenes, far more than S. Typhimurium, which only has 1%. One of these pseudogenes corresponds to sopD2, which in S. Typhimurium encodes an effector protein involved in Salmonella-containing vacuole biogenesis in human epithelial cell lines, which is needed for full virulence of the pathogen. We investigated whether S. Typhi trans-complemented with the functional sopD2 gene from S. Typhimurium (sopD2(STM) ) would reduce the invasion of human epithelial cell lines. Our results showed that the presence of sopD2(STM) in S. Typhi significantly modified the bacterial ability to alter cellular permeability and decrease the CFUs recovered after cell invasion of human epithelial cell line. These results add to mounting evidence that pseudogenes contribute to S. Typhi adaptation to humans. 相似文献
50.
Actinobacillus pleuropneumoniae is the causative agent of a necrotizing hemorrhagic pleuropneumonia in swine. In this study, we investigate the possibility that the limitation of branched-chain amino acids is a stimulus that A. pleuropneumoniae will encounter during infection and will respond to by up-regulation of genes involved in branched-chain amino acid biosynthesis and virulence. Actinobacillus pleuropneumoniae genetic loci that are specifically induced during infection were screened in vitro for expression in response to limitation of branched-chain amino acids. Of 32 in vivo induced promoter clones screened in vitro, eight were induced on chemically defined medium without isoleucine, leucine and valine as compared to complete chemically defined medium. We identify the genomic context of each clone and discuss its relevance to branched-chain amino acid limitation and virulence. We conclude that limitation of branched-chain amino acids is a cue for expression of a subset in vivo induced genes, including not only genes involved in the biosynthesis of branched-chain amino acids, but also other genes that are induced during infection of the natural host. These results suggest that limitation of branched-chain amino acids may be one of an array of environmental cues responsible for the induction of virulence-associated genes in A. pleuropneumoniae. 相似文献