全文获取类型
收费全文 | 2218篇 |
免费 | 162篇 |
国内免费 | 1篇 |
出版年
2024年 | 1篇 |
2023年 | 11篇 |
2022年 | 31篇 |
2021年 | 57篇 |
2020年 | 17篇 |
2019年 | 43篇 |
2018年 | 81篇 |
2017年 | 50篇 |
2016年 | 71篇 |
2015年 | 105篇 |
2014年 | 128篇 |
2013年 | 173篇 |
2012年 | 190篇 |
2011年 | 207篇 |
2010年 | 133篇 |
2009年 | 112篇 |
2008年 | 123篇 |
2007年 | 167篇 |
2006年 | 143篇 |
2005年 | 120篇 |
2004年 | 110篇 |
2003年 | 111篇 |
2002年 | 83篇 |
2001年 | 24篇 |
2000年 | 6篇 |
1999年 | 12篇 |
1998年 | 13篇 |
1997年 | 5篇 |
1996年 | 3篇 |
1995年 | 8篇 |
1994年 | 2篇 |
1993年 | 6篇 |
1992年 | 5篇 |
1990年 | 2篇 |
1988年 | 4篇 |
1987年 | 6篇 |
1986年 | 2篇 |
1985年 | 1篇 |
1984年 | 2篇 |
1983年 | 2篇 |
1981年 | 1篇 |
1976年 | 1篇 |
1975年 | 1篇 |
1973年 | 1篇 |
1970年 | 1篇 |
1968年 | 6篇 |
排序方式: 共有2381条查询结果,搜索用时 31 毫秒
101.
Sarnataro D Paladino S Campana V Grassi J Nitsch L Zurzolo C 《Traffic (Copenhagen, Denmark)》2002,3(11):810-821
PrP(C) is a glycosylphosphatidylinositol-anchored protein expressed in neurons as well as in the cells of several peripheral tissues. Although the normal function of PrP(C) remains unknown, a conformational isoform called PrP(Sc) (scrapie) has been proposed to be the infectious agent of transmissible spongiform encephalopathies in animals and humans. Where and how the PrP(C) to PrP(Sc) conversion occurs in the cells is not yet known. Therefore, dissecting the intracellular trafficking of the wild-type prion protein, as well as of the scrapie isoform, can be of major relevance to the pathogenesis of the diseases. In this report we have analyzed the exocytic pathway of transfected mouse PrP(C) in thyroid and kidney polarized epithelial cells. In contrast to the majority of glycosylphosphatidylinositol-anchored proteins, we found that PrP(C) is localized mainly on the basolateral domain of the plasma membrane of both cell lines. This is reminiscent of the predominant somatodendritic localization found in neurons. However, similarly to apical glycosylphosphatidylinositol-proteins, PrP(C) associates with detergent-resistant microdomains, which have been suggested to have a role in apical sorting of glycosylphosphatidylinositol-proteins, as well as in the conversion process of PrP(C) to PrP(Sc). In order to discriminate whether detergent-resistant microdomains have a direct role in PrP(Sc) conversion, or whether they are involved in the transport of the protein to the site of its conversion, we have examined the effect of disruption of detergent-resistant microdomain association on PrP(C) intracellular traffic. Consistent with the unusual basolateral localization of this glycosylphosphatidylinositol-linked protein, our data exclude a classical role for detergent-resistant microdomains in the post-trans-Golgi network sorting and transport of PrP(C) to the plasma membrane. 相似文献
102.
103.
DNA-based methods for the detection and the identification of phytoplasmas in insect vector extracts
DNA extraction and storage methods have been evaluated with laboratory-reared leafhoppers and/or field-collected leafhoppers
and psyllids. Detection of four different phytopathogenic phytoplasmas, belonging to three taxonomic groups, has been achieved
by several direct or nested polymerase chain reaction (PCR) methods with such DNA extracts. Reactions differed in both the
16/23S ribosomal primer pairs used and the specific assay and cycling conditions. Merits and possible hindrances of the various
primer pairs, in relation to insect DNA extracts, are discussed. However, identification of the phytoplasma(s) necessarily
relied on comparison of the polymorphism in length of the amplified DNA fragments obtained by restriction with appropriate
endonucleases. Endonuclease digestion is crucial for determining the identity (subgroup affiliation) of phytoplasmas of the
same groups that can be carried by an individual vector. 相似文献
104.
105.
Three-hundred twenty five droppings from parrots raised in the premises of 4 breeders and in several private households were
cultured for yeasts. One-hundred sixty droppings (49.2%) resulted positive. From these specimens 212 isolates belonging to
27 different species were obtained. Mainly Candida species such as C. albicans, C. catenulata, C. curvata, C. famata, C. glabrata, C. guilliermondi, C. holmii, C. intermedia, C. krusei, C. lambica, C. lusitaniae, C. membranaefaciens, C. parapsilosis, C. pelliculosa, C. sake and C. valida were isolated. Debaryomyces marama, D. polymorphus, Geotrichum sp., Pichia etchelsii, P. ohmeri, Rhodotorula glutinis, R. rubra, Rhodotorula sp., Saccharomyces cerevisiae, S. kluyiveri and Zygosaccharomyces sp. were also obtained. Dark colonies on Staib medium were never observed. The psittacine birds apparently serve as carriers
for several Candida species or their perfect states and to a lesser extent for other opportunistic yeasts such as Rhodotorula, Trichosporon and Saccharomyces spp., which are considered part of the transient microbiota of the gastrointestinal tract. The most striking finding was
the absence of Cryptococcus spp. among the isolates. The present survey confirms the role of pet birds in carrying potential zoonotic yeasts.
This revised version was published online in June 2006 with corrections to the Cover Date. 相似文献
106.
Bettati S Campanini B Vaccari S Mozzarelli A Schianchi G Hazlett TL Gratton E Benci S 《Biochimica et biophysica acta》2002,1596(1):47-54
Proteins utilizing pyridoxal 5'-phosphate as a coenzyme constitute a large superfamily and are currently classified into three functional groups and five structural fold types. Despite the variability of sequences and catalyzed reactions, they share relevant structural, dynamic and functional properties. Therefore, they constitute an optimal system to investigate the relative influence of primary sequence and coenzyme interactions on folding pathways, structural stability and enzymatic function. O-Acetylserine sulfhydrylase is a dimeric pyridoxal 5'-phosphate dependent enzyme that catalyzes the synthesis of L-cysteine from O-acetylserine and sulfide. The time-resolved fluorescence study of O-acetylserine sulfhydrylase unfolding, here reported, indicates that the coenzyme stabilizes the protein structure. The dependence on denaturant concentration of tryptophan lifetimes in the holo- and apo-enzyme demonstrates that the interactions with the coenzyme stabilize the C-terminal domain to a higher extent with respect to the N-terminal domain. This result is discussed in terms of a linkage between the differential stabilization brought about by the coenzyme and the different degrees of conformational flexibility required by the specialized functional role of distinct protein regions. 相似文献
107.
Borrello MG Mercalli E Perego C Degl'Innocenti D Ghizzoni S Arighi E Eroini B Rizzetti MG Pierotti MA 《Biochemical and biophysical research communications》2002,296(3):515-522
The receptor tyrosine kinase RET, with a known role in embryonic development and in human pathologies, is alternatively spliced to yield at least two functional isoforms, which differ only in their carboxyl terminal. Enigma protein is a member of the PDZ-LIM family and is known to interact with the short isoform of RET/PTC2, a chimeric oncoprotein isolated from papillary thyroid carcinoma. Here, we show that Enigma also interacts in intact cells with the short isoform of RET-wt and of its pathologic mutants associated to MEN2 syndromes, RET-C634R and RET-M918T. In contrast, Enigma binds all the corresponding RET long isoforms very poorly and colocalizes with short but not long RET/PTC2 isoforms. The RET docking tyrosine for Enigma is the last but one before the divergence between the two isoforms and we demonstrated that short-isoform-specific amino acid residues +2 to +4 to this tyrosine are required for the interaction of RET/PTC2 with Enigma. 相似文献
108.
Meyer G Bazzini C Bottà G Garavaglia ML Simona R Manfredi R Sironi C De Biasi S Paulmichl M 《Biochemical and biophysical research communications》2002,290(5):1564-1572
In guinea pig gallbladder epithelial cells, an increase in intracellular cAMP levels elicits the rise of anion channel activity. We investigated by patch-clamp techniques whether K(+) channels were also activated. In a cell-attached configuration and in the presence of theophylline and forskolin or 8-Br-cAMP in the cellular incubation bath, an increase of the open probability (P(o)) values for Ca(2+)-activated K(+) channels with a single-channel conductance of about 160 pS, for inward current, was observed. The increase in P(o) of these channels was also seen in an inside-out configuration and in the presence of PKA, ATP, and cAMP, but not with cAMP alone; phosphorylation did not influence single-channel conductance. In the inside-out configuration, the opioid loperamide (10(-5) M) was able to reduce P(o) when it was present either in the microelectrode filling solution or on the cytoplasmic side. Detection in the epithelial cells by RT-PCR of the mRNA corresponding to the alpha subunit of large-conductance Ca(2+)-activated K(+) channels (BK(Ca)) indicates that this gallbladder channel could belong to the BK family. Immunohistochemistry experiments confirm that these cells express the BK alpha subunit, which is located on the apical membrane. Other K(+) channels with lower conductance (40 pS) were not activated either by 8-Br-cAMP (cell-attached) or by PKA + ATP + cAMP (inside-out). These channels were insensitive to TEA(+) and loperamide. The data demonstrate that under conditions that induce secretion, phosphorylation activates anion channels as well as Ca(2+)-dependent, loperamide-sensitive K(+) channels present on the apical membrane. 相似文献
109.
Evidence is provided for a close link between glutamate (Glu) synthesis and the production of reducing power by the oxidative pentose phosphate pathway (OPPP) in barley ( Hordeum vulgare L. var. Alfeo) root plastids. A rapid procedure for isolating organelles gave yields of plastids of over 30%, 60% of which were intact. The formation of Glu by intact plastids fed with glutamine and 2-oxoglutarate, both substrates of glutamate synthase (GOGAT), depends on glucose-6-phosphate (Glc-6-P) supply. The whole process exhibited an apparent K(m Glc-6-P) of 0.45 mM and is abolished by azaserine, a specific inhibitor of GOGAT; ATP caused a decrease in the rate of Glu formation. Glucose and other sugar phosphates were not as effective in supporting Glu synthesis with respect to Glc-6-P; only ribose-5-phosphate, an intermediate of OPPP, supported rates equivalent to Glc-6-P. Glucose-6-phosphate dehydrogenase (Glc6PDH) rapidly purified from root plastids showed an apparent K(m Glc-6-P) of 0.96 mM and an apparent K(m NADP)(+) of 9 micro M. The enzyme demonstrated high tolerance to NADPH, exhibiting a K(i) (NADPH) of 58.6 micro M and selectively reacted with antibodies against potato plastidic, but not chloroplastic, Glc6PDH isoform. The data support the hypothesis that plastidic OPPP is the main site of reducing power supply for GOGAT within the plastids, and suggest that the plastidic OPPP would be able to sustain Glu synthesis under high NADPH:NADP(+) ratios even if the plastidic Glc6PDH may not be functioning at its highest rates. 相似文献
110.
It was previously shown that in rat thyroid PC-Cl3 cell line, a purinergic P2Y receptor increases the concentration of free cytosolic Ca(2+) ([Ca(2+)](i)) via phospholipase C activation. We here studied whether in a transformed cell line (PC-E1Araf) derived from parental PC-Cl3 cells, ATP is still able to transduce the [Ca(2+)](i)-based intracellular signal.We demonstrate the expression of mRNA for P2Y2 in both PC-Cl3 and PC-E1Araf cells; mRNAs for P2Y1, P2Y4, P2Y6 and P2Y11 were absent. In both cell lines activation of P2Y2 receptor provokes a transient increase in [Ca(2+)](i) followed by a lower sustained phase persisting for over 5min in PC-Cl3 and only 1.5 min in PC-E1Araf cells. In both cell lines the [Ca(2+)](i) reached a plateau level significantly higher than the basal [Ca(2+)](i) level persisting for over 10 min. Removal of extracellular Ca(2+) reduced the initial transient response to ATP in PC-Cl3, but not in PC-E1Araf cells, and completely abolished the plateau phase in both cell lines.In the presence of extracellular Ca(2+) thapsigargin (TG) caused a rise in [Ca(2+)](i) significantly higher in PC-Cl3 than transformed PC-E1Araf cells, while in Ca(2+)-free medium the effect of TG was similar in both cell lines. The capacitative Ca(2+)-entry in PC-Cl3 resulted significantly higher than in PC-E1Araf cells.Further studies were performed in order to investigate whether the different effects of ATP on [Ca(2+)](i) was due to variation in divalent cation plasma membrane permeability. PC-E1Araf cells showed a much lower permeability to Ca(2+), Ba(2+), Sr(2+), Mn(2+), and Co(2+) that may be responsible for the differences in purinergic Ca(2+) signaling pathway with respect to parental PC-Cl3 cells. 相似文献