全文获取类型
收费全文 | 17758篇 |
免费 | 1518篇 |
国内免费 | 8篇 |
出版年
2023年 | 90篇 |
2022年 | 189篇 |
2021年 | 396篇 |
2020年 | 225篇 |
2019年 | 278篇 |
2018年 | 356篇 |
2017年 | 309篇 |
2016年 | 488篇 |
2015年 | 824篇 |
2014年 | 911篇 |
2013年 | 1075篇 |
2012年 | 1440篇 |
2011年 | 1315篇 |
2010年 | 878篇 |
2009年 | 837篇 |
2008年 | 1093篇 |
2007年 | 1109篇 |
2006年 | 984篇 |
2005年 | 1025篇 |
2004年 | 930篇 |
2003年 | 857篇 |
2002年 | 828篇 |
2001年 | 150篇 |
2000年 | 129篇 |
1999年 | 187篇 |
1998年 | 239篇 |
1997年 | 164篇 |
1996年 | 142篇 |
1995年 | 110篇 |
1994年 | 121篇 |
1993年 | 119篇 |
1992年 | 103篇 |
1991年 | 84篇 |
1990年 | 119篇 |
1989年 | 99篇 |
1988年 | 84篇 |
1987年 | 88篇 |
1986年 | 49篇 |
1985年 | 81篇 |
1984年 | 91篇 |
1983年 | 59篇 |
1982年 | 79篇 |
1981年 | 67篇 |
1980年 | 49篇 |
1979年 | 46篇 |
1978年 | 37篇 |
1977年 | 34篇 |
1976年 | 30篇 |
1975年 | 20篇 |
1974年 | 28篇 |
排序方式: 共有10000条查询结果,搜索用时 31 毫秒
961.
Swanson PC 《Molecular and cellular biology》2002,22(5):1340-1351
Two lymphoid cell-specific proteins, RAG-1 and RAG-2, initiate V(D)J recombination by introducing DNA breaks at recombination signal sequences (RSSs). Although the RAG proteins themselves bind and cleave DNA substrates containing either a 12-RSS or a 23-RSS, DNA-bending proteins HMG-1 and HMG-2 are known to promote these processes, particularly with 23-RSS substrates. Using in-gel cleavage assays and DNA footprinting techniques, I analyzed the catalytic activity and protein-DNA contacts in discrete 12-RSS and 23-RSS complexes containing the RAG proteins and either HMG-1 or HMG-2. I found that both the cleavage activity and the pattern of protein-DNA contacts in RAG-HMG complexes assembled on 12-RSS substrates closely resembled those obtained from analogous 12-RSS complexes lacking HMG protein. In contrast, 23-RSS complexes containing both RAG proteins and either HMG-1 or HMG-2 exhibited enhanced cleavage activity and displayed an altered distribution of cleavage products compared to 23-RSS complexes containing only RAG-1 and RAG-2. Moreover, HMG-dependent heptamer contacts in 23-RSS complexes were observed. The protein-DNA contacts in RAG-RSS-HMG complexes assembled on 12-RSS or 23-RSS substrates were strikingly similar at comparable positions, suggesting that the RAG proteins mediate HMG-dependent heptamer contacts in 23-RSS complexes. Results of ethylation interference experiments suggest that the HMG protein is positioned 5' of the nonamer in 23-RSS complexes, interacting largely with the side of the duplex opposite the one contacting the RAG proteins. Thus, HMG protein plays the dual role of bringing critical elements of the 23-RSS heptamer into the same phase as the 12-RSS to promote RAG binding and assisting in the catalysis of 23-RSS cleavage. 相似文献
962.
Fertilities varying in time in an unpredictable manner raise the question of the maintenance of polymorphism, and the subsequent
question of the minimal total number of generations spent by the system in impoverished polymorphism. In the one-locus two-allele
model, level surfaces of this minimal time out of a given set K of constraints defining polymorphism are delineated. The surface of null minimal time is also the largest set of genotype
frequencies from which there exists at least one route remaining in K forever. The influence of the range of admissible fertilities clarifies the trade-off between homozygotes and heterozygotes.
Notably, ranges of fertility forbidding a system leaving K to ever return to it are determined. Generally, keeping a chance to regain polymorphism demands a sufficiently relatively
high fertility for matings involving heterozygotes.
Received: 5 January 2001 / Revised version: 20 October 2001 / Published online: 17 May 2002 相似文献
963.
Engineered viruses to select genes encoding secreted and membrane-bound proteins in mammalian cells 总被引:3,自引:1,他引:2 下载免费PDF全文
Moffatt P Salois P Gaumond MH St-Amant N Godin E Lanctôt C 《Nucleic acids research》2002,30(19):4285-4294
We have developed a functional genomics tool to identify the subset of cDNAs encoding secreted and membrane-bound proteins within a library (the ‘secretome’). A Sindbis virus replicon was engineered such that the envelope protein precursor no longer enters the secretory pathway. cDNA fragments were fused to the mutant precursor and expression screened for their ability to restore membrane localization of envelope proteins. In this way, recombinant replicons were released within infectious viral particles only if the cDNA fragment they contain encodes a secretory signal. By using engineered viral replicons to selectively export cDNAs of interest in the culture medium, the methodology reported here efficiently filters genetic information in mammalian cells without the need to select individual clones. This adaptation of the ‘signal trap’ strategy is highly sensitive (1/200 000) and efficient. Indeed, of the 2546 inserts that were retrieved after screening various libraries, more than 97% contained a putative signal peptide. These 2473 clones encoded 419 unique cDNAs, of which 77% were previously annotated. Of the 94 cDNAs encoding proteins of unknown function, 24% either had no match in databases or contained a secretory signal that could not be predicted from electronic data. 相似文献
964.
Detection and quantification of mitochondrial DNA deletions in individual cells by real-time PCR 总被引:8,自引:0,他引:8
He L Chinnery PF Durham SE Blakely EL Wardell TM Borthwick GM Taylor RW Turnbull DM 《Nucleic acids research》2002,30(14):e68
Defects of mitochondrial DNA (mtDNA) are an important cause of disease and play a role in the ageing process. There are multiple copies of the mitochondrial genome in a single cell. In many patients with acquired or inherited mtDNA mutations, there exists a mixture of mutated and wild type genomes (termed heteroplasmy) within individual cells. As a biochemical and clinical defect is only observed when there are high levels of mutated mtDNA, a crucial investigation is to determine the level of heteroplasmic mutations within tissues and individual cells. We have developed an assay to determine the relative amount of deleted mtDNA using real-time fluorescence PCR. This assay detects the vast majority of deleted molecules, thus eliminating the need to develop specific probes. We have demonstrated an excellent correlation with other techniques (Southern blotting and three- primer competitive PCR), and have shown this technique to be sensitive to quantify the level of deleted mtDNA molecules in individual cells. Finally, we have used this assay to investigate patients with mitochondrial disease and shown in individual skeletal muscle fibres that there exist different patterns of abnormalities between patients with single or multiple mtDNA deletions. We believe that this technique has significant advantages over other methods to quantify deleted mtDNA and, employed alongside our method to sequence the mitochondrial genome from single cells, will further our understanding of the role of mtDNA mutations in human disease and ageing. 相似文献
965.
To test the hypothesis that variation in the putative prostate cancer susceptibility gene ELAC2 contributes to the elevated risk of prostate cancer in Afro-Caribbean males from Tobago, we genotyped the S217L and A514T polymorphisms, previously reported to be associated with prostate cancer risk in a large sample of cases and controls. The frequency of the high-risk Leu allele at the S217L site was the same in cases and controls. Both cases and controls were homozygous for the low-risk Ala allele at the A514T site. In addition, we sequenced the exons and 3'- and 5'-flanking regions of ELAC2 in 24 individuals with histologically confirmed prostate cancer. We identified 17 new single nucleotide polymorphisms. An A(-1196)T polymorphism, which alters a predicted TATA box consensus sequence, was tested in cases and controls, and no significant difference in allele or genotype frequencies was observed. The absence of ELAC2 mutations and lack of association between polymorphisms in ELAC2 and prostate cancer in cases and controls leads us to conclude that ELAC2 does not contribute significantly to the elevated prevalence of prostate cancer in Afro-Caribbean males of Tobago. 相似文献
966.
MacDonald PE Wang G Tsuk S Dodo C Kang Y Tang L Wheeler MB Cattral MS Lakey JR Salapatek AM Lotan I Gaisano HY 《Molecular endocrinology (Baltimore, Md.)》2002,16(11):2452-2461
Insulin secretion is initiated by ionic events involving membrane depolarization and Ca(2+) entry, whereas exocytic SNARE (soluble N-ethylmaleimide-sensitive factor attachment protein receptor) proteins mediate exocytosis itself. In the present study, we characterize the interaction of the SNARE protein SNAP-25 (synaptosome-associated protein of 25 kDa) with the beta-cell voltage-dependent K(+) channel Kv2.1. Expression of Kv2.1, SNAP-25, and syntaxin 1A was detected in human islet lysates by Western blot, and coimmunoprecipitation studies showed that heterologously expressed SNAP-25 and syntaxin 1A associate with Kv2.1. SNAP-25 reduced currents from recombinant Kv2.1 channels by approximately 70% without affecting channel localization. This inhibitory effect could be partially alleviated by codialysis of a Kv2.1N-terminal peptide that can bind in vitro SNAP-25, but not the Kv2.1C-terminal peptide. Similarly, SNAP-25 blocked voltage-dependent outward K(+) currents from rat beta-cells by approximately 40%, an effect that was completely reversed by codialysis of the Kv2.1N fragment. Finally, SNAP-25 had no effect on outward K(+) currents in beta-cells where Kv2.1 channels had been functionally knocked out using a dominant-negative approach, indicating that the interaction is specific to Kv2.1 channels as compared with other beta-cell Kv channels. This study demonstrates that SNAP-25 can regulate Kv2.1 through an interaction at the channel N terminus and supports the hypothesis that SNARE proteins modulate secretion through their involvement in regulation of membrane ion channels in addition to exocytic membrane fusion. 相似文献
967.
968.
Langer I Vertongen P Perret J Waelbroeck M Robberecht P 《Molecular endocrinology (Baltimore, Md.)》2002,16(5):1089-1096
The stimulatory effect of VIP on intracellular calcium concentration ([Ca(2+)](i)) has been investigated in Chinese hamster ovary cells stably transfected with the reporter gene aequorin, and expressing human VPAC(1), VPAC(2), chimeric VPAC(1)/VPAC(2), or mutated receptors. The VIP-induced [Ca(2+)](i) increase was linearly correlated with receptor density and was higher in cells expressing VPAC(1) receptors than in cells expressing a similar VPAC(2) receptor density. The study was performed to establish the receptor sequence responsible for that difference. VPAC(1)/VPAC(2) chimeric receptors were first used for a broad positioning: those having the third intracellular loop (IC(3)) of the VPAC(1) or of the VPAC(2) receptor behaved, in that respect, phenotypically like VPAC(1) and VPAC(2) receptor, respectively. Replacement in the VPAC(2) receptor of the sequence 315-318 (VGGN) within the IC(3) by its VPAC(1) receptor counterpart 328-331 (IRKS) and the introduction of VGGN in state of IRKS in VPAC(1) was sufficient to mimic the VPAC(1) and VPAC(2) receptor characteristics, respectively. Thus, a small sequence in the IC(3) of the VPAC(1) receptor, probably through interaction with G(alphai) and G(alphaq) proteins, is responsible for the efficient agonist-stimulated [Ca(2+)](i) increase. 相似文献
969.
A rapid method to capture and screen for transcription factors by SELDI mass spectrometry. 总被引:12,自引:0,他引:12
970.
Cellular localisation of a water-soluble fullerene derivative 总被引:6,自引:0,他引:6
Foley S Crowley C Smaihi M Bonfils C Erlanger BF Seta P Larroque C 《Biochemical and biophysical research communications》2002,294(1):116-119
Fullerenes are a new class of compounds with potential uses in biology and medicine and many insights were made in the knowledge of their interaction with various biological systems. However, their interaction with organised living systems as well as the site of their potential action remains unclear. In this work, we have demonstrated that a fullerene derivative could cross the external cellular membrane and it localises preferentially to the mitochondria. We propose that our finding supports the potential use of fullerenes as drug delivery agents as their structure mimics that of clathrin known to mediate endocytosis. 相似文献