全文获取类型
收费全文 | 5851篇 |
免费 | 553篇 |
出版年
2023年 | 39篇 |
2022年 | 36篇 |
2021年 | 174篇 |
2020年 | 125篇 |
2019年 | 132篇 |
2018年 | 125篇 |
2017年 | 102篇 |
2016年 | 188篇 |
2015年 | 358篇 |
2014年 | 364篇 |
2013年 | 354篇 |
2012年 | 489篇 |
2011年 | 471篇 |
2010年 | 266篇 |
2009年 | 203篇 |
2008年 | 270篇 |
2007年 | 284篇 |
2006年 | 257篇 |
2005年 | 222篇 |
2004年 | 160篇 |
2003年 | 154篇 |
2002年 | 172篇 |
2001年 | 124篇 |
2000年 | 106篇 |
1999年 | 73篇 |
1998年 | 52篇 |
1997年 | 36篇 |
1996年 | 27篇 |
1995年 | 23篇 |
1993年 | 40篇 |
1992年 | 59篇 |
1991年 | 58篇 |
1990年 | 51篇 |
1989年 | 55篇 |
1988年 | 55篇 |
1987年 | 50篇 |
1986年 | 49篇 |
1985年 | 38篇 |
1984年 | 38篇 |
1983年 | 29篇 |
1982年 | 24篇 |
1979年 | 34篇 |
1978年 | 31篇 |
1977年 | 31篇 |
1975年 | 33篇 |
1974年 | 33篇 |
1973年 | 38篇 |
1972年 | 28篇 |
1971年 | 22篇 |
1969年 | 25篇 |
排序方式: 共有6404条查询结果,搜索用时 125 毫秒
31.
A C-terminal domain of GAP is sufficient to stimulate ras p21 GTPase activity. 总被引:41,自引:8,他引:33 下载免费PDF全文
M S Marshall W S Hill A S Ng U S Vogel M D Schaber E M Scolnick R A Dixon I S Sigal J B Gibbs 《The EMBO journal》1989,8(4):1105-1110
The cDNA for bovine ras p21 GTPase activating protein (GAP) has been cloned and the 1044 amino acid polypeptide encoded by the clone has been shown to bind the GTP complexes of both normal and oncogenic Harvey (Ha) ras p21. To identify the regions of GAP critical for the catalytic stimulation of ras p21 GTPase activity, a series of truncated forms of GAP protein were expressed in Escherichia coli. The C-terminal 343 amino acids of GAP (residues 702-1044) were observed to bind Ha ras p21-GTP and stimulate Ha ras p21 GTPase activity with the same efficiency (kcat/KM congruent to 1 x 10(6) M-1 s-1 at 24 degrees C) as GAP purified from bovine brain or full-length GAP expressed in E. coli. Deletion of the final 61 amino acid residues of GAP (residues 986-1044) rendered the protein insoluble upon expression in E. coli. These results define a distinct catalytic domain at the C terminus of GAP. In addition, GAP contains amino acid similarity with the B and C box domains conserved among phospholipase C-II, the crk oncogene product, and the non-receptor tyrosine kinase oncogene products. This homologous region is located in the N-terminal half of GAP outside of the catalytic domain that stimulates ras p21 GTPase activity and may constitute a distinct structural or functional domain within the GAP protein. 相似文献
32.
33.
Sequence Analysis of N-Ethyl-N-Nitrosourea-Induced Vermilion Mutations in Drosophila Melanogaster 总被引:5,自引:5,他引:0 下载免费PDF全文
The mutational specificity of N-ethyl-N-nitrosourea (ENU) was determined in Drosophila melanogaster using the vermilion locus as a target gene. 25 mutants (16 F1 and 9 F2 mutants) were cloned and sequenced. Only base-pair changes were observed; three of the mutants represented double base substitutions. Transition mutations were the most prominent sequence change: 61% were GC----AT and 18% AT----GC substitutions. Both sequence changes can be explained by the miscoding properties of the modified guanine and thymine bases. A strong bias of neighboring bases on the occurrence of the GC----AT transitions or a strand preference of both types of transition mutations was not observed. The spectrum of ENU mutations in D. melanogaster includes a significant fraction (21%) of transversion mutations. Our data indicate that like in other prokaryotic and eukaryotic systems also in D. melanogaster the O6-ethylguanine adduct is the most prominent premutational lesion after ENU treatment. The strong contribution of the O6-ethylguanine adduct to the mutagenicity of ENU possibly explains the absence of distinct difference between the type of mutations observed in the F1 and F2 mutants. Although the latter arise later during development, the spectrum of mosaic mutations is also dominated by GC----AT transition mutations. 相似文献
34.
Characterization of a parallel-stranded DNA hairpin 总被引:3,自引:0,他引:3
Recently we have shown that synthetic DNA containing homooligomeric A-T base pairs can form a parallel-stranded intramolecular hairpin structure [van de Sande et al. (1988) Science (Washington, D.C.) 241, 551-557]. In the present study, we have employed NMR and optical spectroscopy to investigate the structure of the parallel-stranded (PS) DNA hairpin 3'-d(T)8C4(A)8-3' and the related antiparallel (APS) hairpin 5'-d(T)8C4(A)8-3'. The parallel orientation of the strands in the PS oligonucleotide is achieved by introducing a 5'-5' phosphodiester linkage in the hairpin loop. Ultraviolet spectroscopic and fluorescence data of drug binding are consistent with the formation of PS and APS structures, respectively, in these two hairpins. Vacuum circular dichroism measurements in combination with theoretical CD calculations indicate that the PS structure forms a right-handed helix. 31P NMR measurements indicate that the conformation of the phosphodiester backbone of the PS structure is not drastically different from that of the APS control. The presence of slowly exchanging imino protons at 14 ppm and the observation of nuclear Overhauser enhancement between imino protons and the AH-2 protons demonstrate that similar base pairing and base stacking between T and A residues occur in both hairpins. However, the small chemical shift dispersion observed in proton NMR spectra of the PS hairpin suggests that the stem of this hairpin is more regular than that of the APS hairpin. On the basis of NOESY measurements, we find that the orientation of the bases is in the anti region and that the sugar puckering is in the 2'-endo range.(ABSTRACT TRUNCATED AT 250 WORDS) 相似文献
35.
Perturbation of DNA hairpins containing the EcoRI recognition site by hairpin loops of varying size and composition: physical (NMR and UV) and enzymatic (EcoRI) studies. 总被引:3,自引:2,他引:1 下载免费PDF全文
M W Germann B W Kalisch P Lundberg H J Vogel J H van de Sande 《Nucleic acids research》1990,18(6):1489-1498
We have investigated loop-induced structural perturbation of the stem structure in hairpins d(GAATTCXnGAATTC) (X = A, T and n = 3, 4, 5 and 6) that contain an EcoRI restriction site in close proximity to the hairpin loop. Oligonucleotides containing either a T3 or a A3 loop were not hydrolyzed by the restriction enzyme and also showed only weak binding to EcoRI in the absence of the cofactor Mg2+. In contrast, hairpins with larger loops are hydrolyzed by the enzyme at the scission site next to the loop although the substrate with a A4 loop is significantly more resistant than the oligonucleotide containing a T4 loop. The hairpin structures with 3 loop residues were found to be thermally most stable while larger hairpin loops resulted in structures with lower melting temperatures. The T-loop hairpins are thermally more stable than the hairpins containing the same number of A residues in the loop. As judged from proton NMR spectroscopy and the thermodynamic data, the base pair closest to the hairpin loop did form in all cases studied. The hairpin loops did, however, affect the conformation of the stem structure of the hairpins. From 31P and 1H NMR spectroscopy we conclude that the perturbation of the stem structure is stronger for smaller hairpin loops and that the extent of the perturbation is limited to 2-3 base pairs for hairpins with T3 or A4 loops. Our results demonstrate that hairpin loops modulate the conformation of the stem residues close to the loop and that this in turn reduces the substrate activity for DNA sequence specific proteins. 相似文献
36.
Summary We describe the use of different reporter groups in visualizing replication patterns on metaphase chromosomes after BrdU incorporation by the BrdU antibody technique for the electron microscope. There is an inverse correlation between the density of the label and the size of the reporter particles. This observation alludes to stereo problems interfering with the access of the labeled antibodies into the chromatin. The use of silver enhancement enables easy detection of 1-nm and 5-nm gold particles, which make replication patterns visible in the electron microscope as does the diaminobenzidine/ H2O2 reaction. Possible consequences for the demonstration of replication patterns and for nonradioactive DNA in situ hybridization are discussed. 相似文献
37.
Simultaneous saccharification and protein enrichment fermentation of sugar beet pulp 总被引:1,自引:0,他引:1
Summary A product with 40 % protein content was obtained from sugar beet pulp (1.25–2.0 mm) in 48 h one stage (simultaneous) saccharification/fermentation process under optimized conditions using a specific enzyme mixture andCandida
tropicalis strain, also saving about 40 % enzymes in comparison to a 2-stage process. 相似文献
38.
Summary For the protein upgrading of sugar-beet pulp in solid state fermentation byTrichoderma
reesei andFusarium
oxysporum, serveral conditions were studied to prepare an economical preculture for large scale process. The best performance was shown by a preculture obtained in 24 h from 1.5 % molasses solution at pH 4.5–5.0 with 1.0 % milled beet pulp. 相似文献
39.
Production of tumor necrosis factor by rIFN-gamma-primed C3H/HeJ (Lpsd) macrophages requires the presence of lipid A-associated proteins 总被引:20,自引:0,他引:20
Previous studies have shown that the activation of murine macrophages to a fully tumoricidal state requires that specific environmental signals be delivered to the macrophage in a step-wise manner: a "priming" signal first renders the macrophage stimulated, but not cytolytic. The addition of a second or "trigger" signal to the primed macrophage results in tumoricidal activity. One potent priming signal has been identified as IFN-gamma and one often used trigger signal for endotoxin-responsive (Lpsn) macrophages is LPS. In contrast to LPS-responsive macrophage, rIFN-gamma-primed C3H/HeJ (Lpsd) macrophages fail to become cytolytic in response to protein-free, phenol-water-extracted LPS preparations, but become tumoricidal when exposed in vitro to protein-rich butanol-extracted LPS or purified lipid A-associated proteins. Further characterization of the activation requirements of the C3H/HeJ macrophages revealed that for optimal elaboration of TNF in vitro, two signals were also required: rIFN-gamma and a second signal that contained LAP. C3H/HeJ macrophages macrophages primed with rIFN-gamma failed to produce TNF in response to any concentration of protein-free phenol-water extracted LPS, even when supernatants were concentrated before assaying for functional activity in a standard TNF L929 fibroblast assay. Although exposure of rIFN-gamma-primed C3H/HeJ macrophages to LAP resulted in a fully tumoricidal state equivalent to that exhibited by C3H/OuJ macrophages, the levels of TNF produced remained discrepant. Under identical conditions, C3H/OuJ macrophages produced approximately fivefold more TNF (11,776 U/ml) than C3H/HeJ macrophages (2,399 U/ml). This suggests that although C3H/HeJ macrophages can respond functionally in a "normal" manner given the correct signals, they remain quantitatively deficient in the production of certain proteins. In this system, the elaboration of TNF and macrophage-mediated tumor cell lysis were shown to be dissociable events. The tumor target used in these studies (P815) was shown to be resistant to as much as 40,000 U/ml of purified rTNF. In addition, C3H/OuJ macrophage cultures exposed to LPS only (which resulted in the production of high levels of TNF), failed to lyse these targets. Lastly, anti-mouse TNF antibody added to macrophage cultures had no effect on the induction of tumor cell lysis. 相似文献
40.
The oil-bee/oil-flower relationships: parallelism and other aspects of their evolution in space and time A survey is given of our present knowledge and existing hypotheses concerning the biogeography, history, and phylogeny of plant taxa yielding fatty oil as a floral reward, and of the bee genera involved in their pollination. Four syngenetic complexes of the symbiosis arose convergently: The neotropical, the paleotropical, the holarctic, and the capensic complex. On the basis of the mutual structural adaptations of bees and flowers it is concluded that, in addition, parallelism within related groups as a result of a common tendency to develop the respective organs, has played an important role in the evolution of the oil-based floral interrelationships. 相似文献