全文获取类型
收费全文 | 5537篇 |
免费 | 441篇 |
国内免费 | 7篇 |
出版年
2024年 | 7篇 |
2023年 | 33篇 |
2022年 | 106篇 |
2021年 | 155篇 |
2020年 | 92篇 |
2019年 | 101篇 |
2018年 | 122篇 |
2017年 | 110篇 |
2016年 | 201篇 |
2015年 | 313篇 |
2014年 | 362篇 |
2013年 | 397篇 |
2012年 | 497篇 |
2011年 | 426篇 |
2010年 | 297篇 |
2009年 | 247篇 |
2008年 | 330篇 |
2007年 | 330篇 |
2006年 | 285篇 |
2005年 | 251篇 |
2004年 | 245篇 |
2003年 | 255篇 |
2002年 | 219篇 |
2001年 | 48篇 |
2000年 | 35篇 |
1999年 | 48篇 |
1998年 | 50篇 |
1997年 | 34篇 |
1996年 | 38篇 |
1995年 | 29篇 |
1994年 | 30篇 |
1993年 | 33篇 |
1992年 | 37篇 |
1991年 | 17篇 |
1990年 | 12篇 |
1989年 | 15篇 |
1988年 | 13篇 |
1987年 | 18篇 |
1986年 | 13篇 |
1985年 | 13篇 |
1984年 | 11篇 |
1983年 | 9篇 |
1980年 | 6篇 |
1979年 | 6篇 |
1978年 | 7篇 |
1977年 | 9篇 |
1969年 | 5篇 |
1950年 | 5篇 |
1932年 | 3篇 |
1928年 | 5篇 |
排序方式: 共有5985条查询结果,搜索用时 26 毫秒
51.
Matthieu Eveillard Valérie Fourel Marie-Claude Bare Sophie Kernéis Marie-Hélène Coconnier Tuomo Karjalainen Pierre Bourlioux Alain L. Servin 《Molecular microbiology》1993,7(3):371-381
Experiments reported in this communication showed that the highly toxinogenic Cd 79685, Cd 4784, and Wilkins Clostridium difficile strains and the moderately toxinogenic FD strain grown in the presence of blood adhere to polarized monolayers of two cultured human intestinal cell lines: the human colonic epithelial Caco-2 cells and the human mucus-secreting HT29-MTX cells. Scanning electron microscopy revealed that the bacteria interacted with well-defined apical microvilli of differentiated Caco-2 cells and that the bacteria strongly bind to the mucus layer that entirely covers the surface of the HT29-MTX cells. The binding of C. difficile to Caco-2 cells developed in parallel with the differentiation features of the Caco-2 cells, suggesting that the protein(s) which constitute C. difficile-binding sites are differentiation-related brush border protein(s). To better define this interaction, we tentatively characterized the mechanism(s) of adhesion of C. difficile with adherence assays. It was shown that heating of C. difficile grown in the presence of blood enhanced the bacterial interaction with the brush border of the enterocyte-like Caco-2 cells and the human mucus-secreting HT29-MTX cells. A labile surface-associated component was involved in C. difficile adhesion since washes of C. difficile grown in the presence of blood without heat shock decreased adhesion. After heating, washes of C. difficile grown in the presence of blood did not modify adhesion. Analysis of surface-associated proteins of C. difficile subjected to different culture conditions was con-ducted. After growth of C. difficile Cd 79685, Cd 4784, FD and Wilkins strains in the presence of blood and heating, two predominant SDS-extractable proteins with molecular masses of 12 and 27 kDa were observed and two other proteins with masses of 48 and 31 kDa disappeared. Direct involvement of the 12 and 27 kDa surface-associated proteins in the adhe-sion of C. difficile strains was demonstrated by using rat polycolonal antibodies pAb 12 and pAb 27 directed against the 12 and 27kDa proteins. Indeed, adhesion to Caco-2 cell monoiayers of C. difficiie strains grown in the presence of blood, without or with heat-shock, was blocked. Taken together, our results suggest that C. difficiie may utilize blood components as adhesins to adhere to human intestinal cultured cells. 相似文献
52.
Functional analysis of the cya promoter of Bordetella pertussis 总被引:3,自引:0,他引:3
53.
54.
Tanja Albrecht Sophie Haebel Anke Koch Ulrike Krause Nora Eckermann Martin Steup 《European journal of biochemistry》2004,271(20):3978-3989
Saccharomyces cerevisiae possesses two glycogenin isoforms (designated as Glg1p and Glg2p) that both contain a conserved tyrosine residue, Tyr232. However, Glg2p possesses an additional tyrosine residue, Tyr230 and therefore two potential autoglucosylation sites. Glucosylation of Glg2p was studied using both matrix-assisted laser desorption ionization and electrospray quadrupole time of flight mass spectrometry. Glg2p, carrying a C-terminal (His6) tag, was produced in Escherichia coli and purified. By tryptic digestion and reversed phase chromatography a peptide (residues 219-246 of the complete Glg2p sequence) was isolated that contained 4-25 glucosyl residues. Following incubation of Glg2p with UDPglucose, more than 36 glucosyl residues were covalently bound to this peptide. Using a combination of cyanogen bromide cleavage of the protein backbone, enzymatic hydrolysis of glycosidic bonds and reversed phase chromatography, mono- and diglucosylated peptides having the sequence PNYGYQSSPAM were generated. MS/MS spectra revealed that glucosyl residues were attached to both Tyr232 and Tyr230 within the same peptide. The formation of the highly glucosylated eukaryotic Glg2p did not favour the bacterial glycogen accumulation. Under various experimental conditions Glg2p-producing cells accumulated approximately 30% less glycogen than a control transformed with a Glg2p lacking plasmid. The size distribution of the glycogen and extractable activities of several glycogen-related enzymes were essentially unchanged. As revealed by high performance anion exchange chromatography, the intracellular maltooligosaccharide pattern of the bacterial cells expressing the functional eukaryotic transgene was significantly altered. Thus, the eukaryotic glycogenin appears to be incompatible with the bacterial initiation of glycogen biosynthesis. 相似文献
55.
56.
Marie-Laure Follet-Gueye Sophie Pagny Lo?c Faye Véronique Gomord Azeddine Driouich 《The journal of histochemistry and cytochemistry》2003,51(7):931-940
In plant systems, the green fluorescent protein (GFP) is increasingly used as a marker to study dynamics of the secretory apparatus using fluorescence microscopy. The purpose of this study was to immunogold localize the GFP, at the electron microscopic level, in a line of tobacco BY-2-cultured cells, expressing a GFP-tagged Golgi glycosyltransferase. To this end we have developed a simple, one-step chemical fixation method that allow good structural preservation and specific labeling with anti-GFP antibodies. Using this method, we have been able to show that an N-glycan GFP-tagged xylosyltransferase is specifically associated with Golgi stacks of BY-2 transformed cells and is preferentially located in medial cisternae. As an alternative to cryofixation methods, such as high-pressure freezing, which requires specialized and expensive equipment not available in most laboratories, this method offers researchers the opportunity to investigate GFP-tagged proteins of the endomembrane system in tobacco BY-2 cells. 相似文献
57.
Patrick Jara Sophie Gilbert Pascal Delmas Jean-Claude Guillemot Mourad Kaghad Pascual Ferrara Gérard Loison 《Molecular genetics and genomics : MGG》1996,250(1):97-105
Two new proteinases secreted byCryphonectria parasitica, namely EapB and EapC, have been purified. The corresponding structural genes were isolated by screening a cosmid library, and sequenced. Comparison of genomic and cDNA sequences revealed that theeapB andeapC genes contain three and two introns, respectively. The products of theeapB andeapC genes as deduced from the nucleotide sequences, are 268 and 269 residues long, respectively. N-terminal amino acid sequencing data indicates that EapC is synthesized as a zymogen, which yields a mature 206-amino acid enzyme after cleavage of the prepro sequence. Similarly, sequence alignment studies suggest that EapB is secreted as a 203-residue form which shares extensive similarities not only with EapC but also with two other acid fungal proteinases. However, they display distinct structural features; for example, no cysteine residue is found in EapC. TheeapC gene was mutated using a two-step gene replacement strategy which allowed the specific introduction of several stop codons at the beginning of theeapC coding sequence in an endothiapepsin-deficient (EapA+)C. parasitica strain. Although the resulting strain did not secrete EapC, it still exhibited residual extracellular proteolytic activity, which could be due to EapB. 相似文献
58.
B. Le Rü Sophie Renard Marie-Rose Allo J. Le Lannic J. P. Rolland 《Entomologia Experimentalis et Applicata》1995,77(1):31-36
The ultrastructure of the sensory receptors located on the labium of the cassava mealybugPhenacoccus manihoti Matile-Ferrero (Homoptera, Pseudococcidae) was studied with scanning and transmission electron microscopes. Trichoid hairs
of probable mechanoreceptive function are distributed over the labium. Uniporous chemosensilla which possess a mechanoreceptive
dendrite, multiporous chemosensilla and mechanoreceptive pegs are present on the tip of the labium. The presence of contact
and olfactory chemoreceptors on the labial tip ofP. manihoti suggests that tapping it on the cassava leaf provides the pest with information about the chemical nature of the leaf surface.
ORSTOM 相似文献
59.
J. C. du Preez M. Immelman J. L. F. Kock S. G. Kilian 《World journal of microbiology & biotechnology》1997,13(1):81-87
The effect of different initial acetic acid concentrations on the growth of and lipid and gamma-linolenic acid (GLA) production
byMucor circinelloides CBS 203.28 was determined in a 14 litre stirred tank reactor operated in a fedbatch, pH-stat mode with acetic acid as carbon
source and pH titrant. Increased acetic acid concentrations in the culture resulted in a significant increase in the crude
oil content of the biomass. By contrast, all the other parameters such as the biomass concentration, GLA and oil yield on
acetic acid, the GLA content of the biomass and oil, the growth rate and volumetric rate of GLA production decreased with
an increase in acetic acid concentration. The best results were obtained with acetic acid at 2 g/1, which gave 39.8 mg GLA/g
biomass and 15.6% GLA in the neutral lipid fraction, amounting to 340 mg GLA/1 culture. A decrease in the glyco- and phospho-lipid
fractions during the cultivation coincided with an increase in the neutral lipid fraction. The GLA content of the biomass
remained within rather narrow limits of 3.5% to 4% of the biomass, irrespective of the oil content of the biomass. The fatty
acid profile was not greatly affected by the acetic acid concentration. The hyphae of the fungus were characterized by the
accumulation of large intracellular oil droplets and some septa delimited the hyphae. 相似文献
60.
Ivan Laprevotte Sophie Brouillet Christophe Terzian Alain Hénaut 《Journal of molecular evolution》1997,44(2):214-225
A computer-assisted analysis was made of 24 complete nucleotide sequences selected from the vertebrate retroviruses to represent
the ten viral groups. The conclusions of this analysis extend and strengthen the previously made hypothesis on the Moloney
murine leukemia virus: The evolution of the nucleotide sequence appears to have occurred mainly through at least three overlapping
levels of duplication: (1) The distributions of overrepresented (3–6)-mers are consistent with the universal rule of a trend
toward TG/CT excess and with the persistence of a certain degree of symmetry between the two strands of DNA. This suggests
one or several original tandemly repeated sequences and some inverted duplications. (2) The existence of two general core
consensuses at the level of these (3–6)-mers supports the hypothesis of a common evolutionary origin of vertebrate retroviruses.
Consensuses more specific to certain sequences are compatible with phylogenetic trees established independently. The consensuses
could correspond to intermediary evolutionary stages. (3) Most of the (3–6)-mers with a significantly higher than average
frequency appear to be internally repeated (with monomeric or oligomeric internal iterations) and seem to be at least partly
the cause of the bias observed by other researchers at the level of retroviral nucleotide composition. They suggest a third
evolutionary stage by slippage-like stepwise local duplications.
Received: 3 January 1996 / Accepted: 27 March 1996 相似文献