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61.
Extraction of extracellular polymeric substances (EPS) from anaerobic granular sludges: comparison of chemical and physical extraction protocols 总被引:9,自引:0,他引:9
Paul D’Abzac François Bordas Eric Van Hullebusch Piet N. L. Lens Gilles Guibaud 《Applied microbiology and biotechnology》2010,85(5):1589-1599
The characteristics of the extracellular polymeric substances (EPS) extracted with nine different extraction protocols from
four different types of anaerobic granular sludge were studied. The efficiency of four physical (sonication, heating, cationic
exchange resin (CER), and CER associated with sonication) and four chemical (ethylenediaminetetraacetic acid, ethanol, formaldehyde
combined with heating, or NaOH) EPS extraction methods was compared to a control extraction protocols (i.e., centrifugation).
The nucleic acid content and the protein/polysaccharide ratio of the EPS extracted show that the extraction does not induce
abnormal cellular lysis. Chemical extraction protocols give the highest EPS extraction yields (calculated by the mass ratio
between sludges and EPS dry weight (DW)). Infrared analyses as well as an extraction yield over 100% or organic carbon content
over 1 g g−1 of DW revealed, nevertheless, a carry-over of the chemical extractants into the EPS extracts. The EPS of the anaerobic granular
sludges investigated are predominantly composed of humic-like substances, proteins, and polysaccharides. The EPS content in
each biochemical compound varies depending on the sludge type and extraction technique used. Some extraction techniques lead
to a slightly preferential extraction of some EPS compounds, e.g., CER gives a higher protein yield. 相似文献
62.
Jan Bartacek Isabella Manconi Gerardo Sansone Roberta Murgia Piet N.L. Lens 《Nitric oxide》2010,22(2):101-105
Hydrogen sulfide (H2S) inhibits the last step of the denitrification process, i.e. the reduction of nitrous oxide (N2O) to dinitrogen gas (N2), both in natural environments (marine sediments) and industrial processes (activated sludge, methanogenic sludge, BioDeNOx process). In a previously published study, we showed that the inhibitory effect of sulfide to N2O reduction in mixed microbial communities is reversible and can be counteracted by dosing trace amounts of copper. It remained, however, unclear if this was due to copper sulfide precipitation or a retrofitting of the copper containing N2O-reductase (N2OR). The present study aimed to elucidate the mechanism of the restoration of sulfide-inhibited N2O reducing activity by metal addition to a pure Pseudomonas aeruginosa culture. This was done by using other metals (zinc, cobalt and iron) in comparison with copper. Zinc and cobalt clearly alleviated the sulfide inhibition of N2OR to the same extent as copper and the activity restoration was extremely fast (within 15 min, Fig. 3) for zinc, cobalt and copper. This suggests that the alleviation of the inhibitory effect of sulfide is due to metal sulfide precipitation and thus not exclusively limited to Cu. This work also underlines the importance of metal speciation: supply of iron did not restore the N2OR activity because it was precipitated by the phosphates present in the medium and thus could not precipitate the sulfide. 相似文献
63.
CD4+CD28null T cells in autoimmune disease: pathogenic features and decreased susceptibility to immunoregulation 总被引:1,自引:0,他引:1
Thewissen M Somers V Hellings N Fraussen J Damoiseaux J Stinissen P 《Journal of immunology (Baltimore, Md. : 1950)》2007,179(10):6514-6523
To determine the role of expanded CD4(+)CD28(null) T cells in multiple sclerosis and rheumatoid arthritis pathology, these cells were phenotypically characterized and their Ag reactivity was studied. FACS analysis confirmed that CD4(+)CD28(null) T cells are terminally differentiated effector memory cells. In addition, they express phenotypic markers that indicate their capacity to infiltrate into tissues and cause tissue damage. Whereas no reactivity to the candidate autoantigens myelin basic protein and collagen type II was observed within the CD4(+)CD28(null) T cell subset, CMV reactivity was prominent in four of four HC, four of four rheumatoid arthritis patients, and three of four multiple sclerosis patients. The level of the CMV-induced proliferative response was found to be related to the clonal diversity of the response. Interestingly, our results illustrate that CD4(+)CD28(null) T cells are not susceptible to the suppressive actions of CD4(+)CD25(+) regulatory T cells. In conclusion, this study provides several indications for a role of CD4(+)CD28(null) T cells in autoimmune pathology. CD4(+)CD28(null) T cells display pathogenic features, fill up immunological space, and are less susceptible to regulatory mechanisms. However, based on their low reactivity to the autoantigens tested in this study, CD4(+)CD28(null) T cells most likely do not play a direct autoaggressive role in autoimmune disease. 相似文献
64.
Dumont D Noben JP Moreels M Vanderlocht J Hellings N Vandenabeele F Lambrichts I Stinissen P Robben J 《Journal of neurochemistry》2007,102(2):562-576
Oligodendrocytes are glial cells responsible for the synthesis and maintenance of myelin in the central nervous system (CNS). Oligodendrocytes are vulnerable to damage occurring in a variety of neurological diseases. Understanding oligodendrocyte biology is crucial for the dissemination of de- and remyelination mechanisms. The goal of the present study is the construction of a protein database of mature rat oligodendrocytes. Post-mitotic oligodendrocytes were isolated from mature Wistar rats and subjected to immunocytochemistry. Proteins were extracted and analyzed by means of two-dimensional gel electrophoresis and two-dimensional liquid chromatography, both coupled to mass spectrometry. The combination of the gel-based and gel-free approach resulted in confident identification of a total of 200 proteins. A minority of proteins were identified in both proteomic strategies. The identified proteins represent a variety of functional groups, including novel oligodendrocyte proteins. The results of this study emphasize the power of the applied proteomic strategy to study known or to reveal new proteins and to investigate their regulation in oligodendrocytes in different disease models. 相似文献
65.
The protein that binds to DNA base J in trypanosomatids has features of a thymidine hydroxylase 下载免费PDF全文
Yu Z Genest PA ter Riet B Sweeney K DiPaolo C Kieft R Christodoulou E Perrakis A Simmons JM Hausinger RP van Luenen HG Rigden DJ Sabatini R Borst P 《Nucleic acids research》2007,35(7):2107-2115
Trypanosomatids contain an unusual DNA base J (beta-d-glucosylhydroxymethyluracil), which replaces a fraction of thymine in telomeric and other DNA repeats. To determine the function of base J, we have searched for enzymes that catalyze J biosynthesis. We present evidence that a protein that binds to J in DNA, the J-binding protein 1 (JBP1), may also catalyze the first step in J biosynthesis, the conversion of thymine in DNA into hydroxymethyluracil. We show that JBP1 belongs to the family of Fe(2+) and 2-oxoglutarate-dependent dioxygenases and that replacement of conserved residues putatively involved in Fe(2+) and 2-oxoglutarate-binding inactivates the ability of JBP1 to contribute to J synthesis without affecting its ability to bind to J-DNA. We propose that JBP1 is a thymidine hydroxylase responsible for the local amplification of J inserted by JBP2, another putative thymidine hydroxylase. 相似文献
66.
Inhibition of MDR1 expression with altritol-modified siRNAs 总被引:1,自引:1,他引:0
Fisher M Abramov M Van Aerschot A Xu D Juliano RL Herdewijn P 《Nucleic acids research》2007,35(4):1064-1074
Altritol-modified nucleic acids (ANAs) support RNA-like A-form structures when included in oligonucleotide duplexes. Thus altritol residues seem suitable as candidates for the chemical modification of siRNAs. Here we report that ANA-modified siRNAs targeting the MDR1 gene can exhibit improved efficacy as compared to unmodified controls. This was particularly true of ANA modifications at or near the 3′ end of the sense or antisense strands, while modification at the 5′ end of the antisense strand resulted in complete loss of activity. Multiple ANA modifications within the sense strand were also well tolerated. Duplexes with ANA modifications at appropriate positions in both strands were generally more effective than duplexes with one modified and one unmodified strand. Initial evidence suggests that the loss of activity associated with ANA modification of the 5′-antisense strand may be due to reduced phosphorylation at this site by cellular kinases. Treatment of drug resistant cells with MDR1-targeted siRNAs resulted in reduction of P-glycoprotein (Pgp) expression, parallel reduction in MDR1 message levels, increased accumulation of the Pgp substrate rhodamine 123, and reduced resistance to anti-tumor drugs. Interestingly, the duration of action of some of the ANA-modified siRNAs was substantially greater than that of unmodified controls. These observations suggest that altritol modifications may be helpful in developing siRNAs with enhanced pharmacological effectiveness. 相似文献
67.
Morten Skage Anders Hobæk Štĕpánka Ruthová Barbara Keller Adam Petrusek Jaromír Sed’a Piet Spaak 《Hydrobiologia》2007,594(1):19-32
A collaborative research effort was undertaken to evaluate the robustness of a recently developed genetic tool for species
identification of members in the morphologically variable Daphnia longispina species complex. This genetic method, based on restriction fragment length polymorphism (RFLP) of the internal transcribed
spacer region (ITS) of nuclear ribosomal DNA (rDNA) with restriction enzymes Mwo I and Sau96 I [Billiones et al., 2004. Hydrobiologia 526: 43–53], was applied to many different European populations. Results were
compared with two or more independently obtained characters (morphology, allozymes, mitochondrial DNA (mtDNA), or cloned rDNA-ITS
sequences). Individuals of most taxa were readily identified, but unexpected ITS-RFLP patterns were found in many individuals
indicated by other markers to be D. galeata or one of its hybrids. Among 43 investigated D. galeata populations (902 specimen analysed by ITS-RFLP), deviant RFLP fragment patterns occurred in 26 (i.e., more than half) of
the populations. The deviant patterns could be attributed to the loss of one single restriction site in the ITS2 region. This
loss made the distinction of D. galeata from other species unreliable, and F1 hybrids could not be identified. Future users should be aware of this shortcoming of
the Billions et al. [2004. Hydrobiologia 526: 43–53] protocol. As a solution to this problem, we present an improved genetic
identification protocol based on a simple double digestion of the rDNA-ITS region with the restriction enzymes BsrB I and EagI. Sequence analyses of rDNA-ITS clones and preliminary testing indicate that the new protocol is unaffected by the rDNA variation
which troubled the Mwo I/Sau96 I protocol. Further, the new protocol identifies all European species of the D. longispina complex, as well as their F1 hybrids. However, a wider screening is required to verify its general utility for all species,
since yet unknown variation may occur.
Guest editor: Piet Spaak
Cladocera: Proceedings of the 7th International Symposium on Cladocera 相似文献
68.
The complete genome sequence of Campylobacter jejuni strain 81116 (NCTC11828) 总被引:1,自引:0,他引:1 下载免费PDF全文
Pearson BM Gaskin DJ Segers RP Wells JM Nuijten PJ van Vliet AH 《Journal of bacteriology》2007,189(22):8402-8403
Campylobacter jejuni is a major human enteric pathogen that displays genetic variability via genomic reorganization and phase variation. This variability can adversely affect the outcomes and reproducibility of experiments. C. jejuni strain 81116 (NCTC11828) has been suggested to be a genetically stable strain (G. Manning, B. Duim, T. Wassenaar, J. A. Wagenaar, A. Ridley, and D. G. Newell, Appl. Environ. Microbiol. 67:1185-1189, 2001), is amenable to genetic manipulation, and is infective for chickens. Here we report the finished annotated genome sequence of C. jejuni strain 81116. 相似文献
69.
Phosphopantetheine adenylyltransferase from Escherichia coli: investigation of the kinetic mechanism and role in regulation of coenzyme A biosynthesis 下载免费PDF全文
Miller JR Ohren J Sarver RW Mueller WT de Dreu P Case H Thanabal V 《Journal of bacteriology》2007,189(22):8196-8205
Phosphopantetheine adenylyltransferase (PPAT) from Escherichia coli is an essential hexameric enzyme that catalyzes the penultimate step in coenzyme A (CoA) biosynthesis and is a target for antibacterial drug discovery. The enzyme utilizes Mg-ATP and phosphopantetheine (PhP) to generate dephospho-CoA (dPCoA) and pyrophosphate. When overexpressed in E. coli, PPAT copurifies with tightly bound CoA, suggesting a feedback inhibitory role for this cofactor. Using an enzyme-coupled assay for the forward-direction reaction (dPCoA-generating) and isothermal titration calorimetry, we investigated the steady-state kinetics and ligand binding properties of PPAT. All substrates and products bind the free enzyme, and product inhibition studies are consistent with a random bi-bi kinetic mechanism. CoA inhibits PPAT and is competitive with ATP, PhP, and dPCoA. Previously published structures of PPAT crystallized at pH 5.0 show half-the-sites reactivity for PhP and dPCoA and full occupancy by ATP and CoA. Ligand-binding studies at pH 8.0 show that ATP, PhP, dPCoA, and CoA occupy all six monomers of the PPAT hexamer, although CoA exhibits two thermodynamically distinct binding modes. These results suggest that the half-the-sites reactivity observed in PPAT crystal structures may be pH dependent. In light of previous studies on the regulation of CoA biosynthesis, the PPAT kinetic and ligand binding data suggest that intracellular PhP concentrations modulate the distribution of PPAT monomers between high- and low-affinity CoA binding modes. This model is consistent with PPAT serving as a “backup” regulator of pathway flux relative to pantothenate kinase. 相似文献
70.
Genome sequencing and analysis of the versatile cell factory Aspergillus niger CBS 513.88 总被引:1,自引:0,他引:1
Pel HJ de Winde JH Archer DB Dyer PS Hofmann G Schaap PJ Turner G de Vries RP Albang R Albermann K Andersen MR Bendtsen JD Benen JA van den Berg M Breestraat S Caddick MX Contreras R Cornell M Coutinho PM Danchin EG Debets AJ Dekker P van Dijck PW van Dijk A Dijkhuizen L Driessen AJ d'Enfert C Geysens S Goosen C Groot GS de Groot PW Guillemette T Henrissat B Herweijer M van den Hombergh JP van den Hondel CA van der Heijden RT van der Kaaij RM Klis FM Kools HJ Kubicek CP van Kuyk PA Lauber J 《Nature biotechnology》2007,25(2):221-231