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排序方式: 共有216条查询结果,搜索用时 15 毫秒
1.
Yasutaka Kakui Tomonari Sunaga Kunio Arai James Dodgson Liang Ji Attila Csikász-Nagy Rafael Carazo-Salas Masamitsu Sato 《Open biology》2015,5(6)
Integration of an external gene into a fission yeast chromosome is useful to investigate the effect of the gene product. An easy way to knock-in a gene construct is use of an integration plasmid, which can be targeted and inserted to a chromosome through homologous recombination. Despite the advantage of integration, construction of integration plasmids is energy- and time-consuming, because there is no systematic library of integration plasmids with various promoters, fluorescent protein tags, terminators and selection markers; therefore, researchers are often forced to make appropriate ones through multiple rounds of cloning procedures. Here, we establish materials and methods to easily construct integration plasmids. We introduce a convenient cloning system based on Golden Gate DNA shuffling, which enables the connection of multiple DNA fragments at once: any kind of promoters and terminators, the gene of interest, in combination with any fluorescent protein tag genes and any selection markers. Each of those DNA fragments, called a ‘module’, can be tandemly ligated in the order we desire in a single reaction, which yields a circular plasmid in a one-step manner. The resulting plasmids can be integrated through standard methods for transformation. Thus, these materials and methods help easy construction of knock-in strains, and this will further increase the value of fission yeast as a model organism. 相似文献
2.
3.
Calcrete aquifers from the Yilgarn region of arid central Western Australia contain an assemblage of obligate groundwater invertebrate species that are each endemic to single aquifers. Fine-scale phylogeographic and population genetic analyses of three sympatric and independently derived species of amphipod (Chiltoniidae) were carried out to determine whether there were common patterns of population genetic structure or evidence for past geographic isolation of populations within a single calcrete aquifer. Genetic diversity in amphipod mitochondrial DNA (cytochrome c oxidase subunit I gene) and allozymes were examined across a 3.5 km2 region of the Sturt Meadows calcrete, which contains a grid of 115 bore holes (=wells). Stygobiont amphipods were found to have high levels of mitochondrial haplotype diversity coupled with low nucleotide diversity. Mitochondrial phylogeographic structuring was found between haplogroups for one of the chiltoniid species, which also showed population structuring for nuclear markers. Signatures of population expansion in two of the three species, match previous findings for diving beetles at the same site, indicating that the system is dynamic. We propose isolation of populations in refugia within the calcrete, followed by expansion events, as the most likely source of intraspecific genetic diversity, due to changes in water level influencing gene flow across the calcrete. 相似文献
4.
Anna CC Aguiar Ananda C Cunha Isabela Penna Ceravolo Regina A Correia Gon?alves Arildo JB Oliveira Antoniana Ursine Krettli 《Memórias do Instituto Oswaldo Cruz》2015,110(7):906-913
Several species of Aspidosperma plants are used to treat diseases in
the tropics, including Aspidosperma ramiflorum, which acts against
leishmaniasis, an activity that is experimentally confirmed. The species, known as
guatambu-yellow, yellow peroba,
coffee-peroba andmatiambu, grows in the Atlantic
Forest of Brazil in the South to the Southeast regions. Through a guided
biofractionation of A. ramiflorum extracts, the plant activity
against Plasmodium falciparum was evaluated in vitro for toxicity
towards human hepatoma G2 cells, normal monkey kidney cells and nonimmortalised human
monocytes isolated from peripheral blood. Six of the seven extracts tested were
active at low doses (half-maximal drug inhibitory concentration < 3.8 µg/mL); the
aqueous extract was inactive. Overall, the plant extracts and the purified compounds
displayed low toxicity in vitro. A nonsoluble extract fraction and one purified
alkaloid isositsirikine (compound 5) displayed high selectivity indexes (SI) (= 56
and 113, respectively), whereas compounds 2 and 3 were toxic (SI < 10). The
structure, activity and low toxicity of isositsirikine in vitro are described here
for the first time in A. ramiflorum, but only the neutral and
precipitate plant fractions were tested for activity, which caused up to 53%
parasitaemia inhibition of Plasmodium berghei in mice with
blood-induced malaria. This plant species is likely to be useful in the further
development of an antimalarial drug, but its pharmacological evaluation is still
required. 相似文献
5.
J Klein J Gonzalez J Duchene L Esposito JP Pradère E Neau C Delage D Calise A Ahluwalia P Carayon JB Pesquero M Bader JP Schanstra JL Bascands 《FASEB journal》2009,23(1):134-142
Renal fibrosis is the common histological feature of advanced glomerular and tubulointerstitial disease leading to end-stage renal disease (ESRD). However, specific antifibrotic therapies to slow down the evolution to ESRD are still absent. Because persistent inflammation is a key event in the development of fibrosis, we hypothesized that the proinflammatory kinin B1 receptor (B1R) could be such a new target. Here we show that, in the unilateral ureteral obstruction model of renal fibrosis, the B1R is overexpressed and that delayed treatment with an orally active nonpeptide B1R antagonist blocks macrophage infiltration, leading to a reversal of the level of renal fibrosis. In vivo bone marrow transplantation studies as well as in vitro studies on renal cells show that part of this antifibrotic mechanism of B1R blockade involves a direct effect on resident renal cells by inhibiting chemokine CCL2 and CCL7 expression. These findings suggest that blocking the B1R is a promising antifibrotic therapy. 相似文献
6.
Background
DNA repair is the general term for the collection of critical mechanisms which repair many forms of DNA damage such as methylation or ionizing radiation. DNA repair has mainly been studied in experimental and clinical situations, and relatively few information-based approaches to new extracting DNA repair knowledge exist. As a first step, automatic detection of DNA repair proteins in genomes via informatics techniques is desirable; however, there are many forms of DNA repair and it is not a straightforward process to identify and classify repair proteins with a single optimal method. We perform a study of the ability of homology and machine learning-based methods to identify and classify DNA repair proteins, as well as scan vertebrate genomes for the presence of novel repair proteins. Combinations of primary sequence polypeptide frequency, secondary structure, and homology information are used as feature information for input to a Support Vector Machine (SVM). 相似文献7.
Majed F. Alghoribi Tarek M. Gibreel Andrew R. Dodgson Scott A. Beatson Mathew Upton 《PloS one》2014,9(7)
Galleria mellonella larvae are an alternative in vivo model for investigating bacterial pathogenicity. Here, we examined the pathogenicity of 71 isolates from five leading uropathogenic E. coli (UPEC) lineages using G. mellonella larvae. Larvae were challenged with a range of inoculum doses to determine the 50% lethal dose (LD50) and for analysis of survival outcome using Kaplan-Meier plots. Virulence was correlated with carriage of a panel of 29 virulence factors (VF). Larvae inoculated with ST69 and ST127 isolates (104 colony-forming units/larvae) showed significantly higher mortality rates than those infected with ST73, ST95 and ST131 isolates, killing 50% of the larvae within 24 hours. Interestingly, ST131 isolates were the least virulent. We observed that ST127 isolates are significantly associated with a higher VF-score than isolates of all other STs tested (P≤0.0001), including ST69 (P<0.02), but one ST127 isolate (strain EC18) was avirulent. Comparative genomic analyses with virulent ST127 strains revealed an IS1 mediated deletion in the O-antigen cluster in strain EC18, which is likely to explain the lack of virulence in the larvae infection model. Virulence in the larvae was not correlated with serotype or phylogenetic group. This study illustrates that G. mellonella are an excellent tool for investigation of the virulence of UPEC strains. The findings also support our suggestion that the incidence of ST127 strains should be monitored, as these isolates have not yet been widely reported, but they clearly have a pathogenic potential greater than that of more widely recognised clones, including ST73, ST95 or ST131. 相似文献
8.
Phylogenetic relationships were determined for 76 partial P-element
sequences from 14 species of the melanogaster species group within the
Drosophila subgenus Sophophora. These results are examined in the context
of the phylogeny of the species from which the sequences were isolated.
Sequences from the P-element family fall into distinct subfamilies, or
clades, which are often characteristic for particular species subgroups.
When examined locally among closely related species, the evolution of P
elements is characterized by vertical transmission, whereby the P-element
phylogeny traces the species phylogeny. On a broader scale, however, the
P-element phylogeny is not congruent with the species phylogeny. One
feature of P-element evolution in the melanogaster group is the presence of
more than one P-element subfamily, differing by as much as 36%, in the
genomes of some species. Thus, P elements from several individual species
are not monophyletic, and a likely explanation for the incongruence between
P-element and species phylogenies is provided by the comparison of
paralogous sequences. In certain instances, horizontal transfer seems to be
a valid alternative explanation for lack of congruence between species and
P-element phylogenies. The canonical P-element subfamily, which represents
the active, autonomous transposable element, is restricted to D.
melanogaster. Thus, its origin clearly lies outside of the melanogaster
species group, consistent with the earlier conclusion of recent horizontal
transfer.
相似文献
9.
Purification, properties and cellular localization of the stereospecific CS2 secondary alkylsulphohydrolase of Comamonas terrigena. 总被引:2,自引:2,他引:0 下载免费PDF全文
The availability of homogeneous samples of the potassium salts of L- and D-octan-2-yl sulphate has enabled the separation of the optically stereospecific CS1 and CS2 secondary alkysulphohydrolases from extracts of cells of Comamonas terrigena. The CS2 enzyme was purified to homogeneity, and an initial study was made of its general properties, specificity, cellular localization and relationship to the CS1 enzyme. The CS2 enzyme has a molecular weight of approx. 250000 and a subunit size of approx. 58000, indicating that the molecule is a tetramer. Under the experimental conditions used the enzyme appears to be specific for (+)-secondary alkyl sulphate esters with the sulphate group at C-2 and with a chain length of at least six carbons. Enzyme activity towards racemic C-2 sulphates increases with increasing chain length up to C10, and there is some indirect evidence to suggest that activity declines when that chain length is exceeded. Other indirect evidence confirms that the CS1 enzyme exhibits similar specificity, except that only (-)-isomers can serve as substrates. Both enzymes are present in broth-grown stationary-phase cells of C. terrigena in approximately equal amounts. 相似文献
10.
Assay of a microsomal marker enzyme: rat liver arylsulphatase C. 总被引:1,自引:0,他引:1