全文获取类型
收费全文 | 1094篇 |
免费 | 82篇 |
专业分类
1176篇 |
出版年
2023年 | 1篇 |
2022年 | 15篇 |
2021年 | 23篇 |
2020年 | 13篇 |
2019年 | 14篇 |
2018年 | 22篇 |
2017年 | 30篇 |
2016年 | 45篇 |
2015年 | 52篇 |
2014年 | 51篇 |
2013年 | 96篇 |
2012年 | 103篇 |
2011年 | 107篇 |
2010年 | 47篇 |
2009年 | 49篇 |
2008年 | 73篇 |
2007年 | 68篇 |
2006年 | 50篇 |
2005年 | 65篇 |
2004年 | 53篇 |
2003年 | 45篇 |
2002年 | 30篇 |
2001年 | 8篇 |
2000年 | 3篇 |
1999年 | 7篇 |
1998年 | 14篇 |
1997年 | 13篇 |
1996年 | 12篇 |
1995年 | 8篇 |
1994年 | 4篇 |
1993年 | 6篇 |
1992年 | 4篇 |
1991年 | 2篇 |
1990年 | 3篇 |
1989年 | 5篇 |
1988年 | 3篇 |
1987年 | 4篇 |
1986年 | 2篇 |
1985年 | 3篇 |
1983年 | 2篇 |
1982年 | 3篇 |
1981年 | 3篇 |
1980年 | 3篇 |
1979年 | 6篇 |
1977年 | 1篇 |
1974年 | 1篇 |
1973年 | 1篇 |
1964年 | 2篇 |
1958年 | 1篇 |
排序方式: 共有1176条查询结果,搜索用时 0 毫秒
991.
Troiano L Ferraresi R Lugli E Nemes E Roat E Nasi M Pinti M Cossarizza A 《Nature protocols》2007,2(11):2719-2727
The analysis of changes in mitochondrial membrane potential (MMP) that can occur during apoptosis provides precious information on the mechanisms and pathways of cell death. For many years, the metachromatic fluorochrome JC-1 (5,5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolcarbocyanine iodide) was used for this purpose. Thanks to new dyes and to the technical improvements recently adopted in several flow cytometers, it is now possible to investigate, along with MMP, a variety of other parameters. Using three sources of excitation and polychromatic flow cytometry, we have developed a protocol that can be applied to cells undergoing apoptosis. In the model of U937 cells incubated with the chemopreventive agent quercetin (3,3',4',5,7-pentahydroxyflavone), we describe the detection at the single cell level of changes in MMP (by JC-1), early apoptosis (exposition of phosphatidylserine on the plasma membrane detected by annexin-V), late apoptosis and secondary necrosis (decreased DNA content by Hoechst 33342 and permeability of the plasma membrane to propidium iodide). The procedure can be completed in less than 2 h. 相似文献
992.
Luigi Menghini Claudio Ferrante Mohamad Fawzi Mahomoodally Lidia Leporini Marcello Locatelli 《Plant biosystems》2019,153(5):633-639
Cynoglossum creticum Mill (Boraginaceae) is used traditionally as a remedy to manage several human ailments. In this context, the present study aimed to perform multiple pharmacological investigations on the hydroalcoholic extracts prepared from Cynoglossum roots and aerial parts (leaves and flowers). We evaluated the antioxidant and enzyme inhibitory (against cholinesterases, α-glucosidase, α-amylase, lipase and tyrosinase) activity of the extracts. The protective effect(s) of the extracts on cardiomyocyte C2C12 and intestinal HCT116 cell lines challenged with hydrogen peroxide (H2O2) was studied. We found that the aerial parts harbored the highest amount of phenolic compounds. Generally, aerial parts showed significant antioxidant and enzyme inhibitory effects. Leaves exhibited the best lipase inhibitory activity (173.15 mgOE/g extract), followed by flowers and roots. The root and aerial extracts were equally able to blunt intracellular H2O2 induced reactive oxygen species production from both C2C12 and HCT116 cell lines. Both cells lines could be treated with scalar concentrations of root and flower extracts in the range 50–300?μg/mL without interferences on cell viability. In conclusion, the present study showed protective effects exerted by Cynoglossum extracts, which could serve as a foundation for the development of pharmaceuticals and nutraceuticals derived from Cynoglossum. 相似文献
993.
Silica columns are among the most used DNA purification systems, allowing a good yield of high-quality nucleic acids without organic extractions. Silica column regeneration protocols reported up to now to remove DNA traces are time-consuming, and their effectiveness on genomic DNA has not been demonstrated. Here we report a very rapid regeneration procedure that ensures no DNA carryover, independent of its size, without impairing column efficiency. The method takes advantage of the improved DNA removal by low concentrations of Triton X-100. 相似文献
994.
Barbara Lazzari Andrea Caprera Alessandro Cestaro Ivan Merelli Marcello Del Corvo Paolo Fontana Luciano Milanesi Riccardo Velasco Alessandra Stella 《BMC plant biology》2009,9(1):1-13
Background
Peanut (Arachis hypogaea L.) is widely used as a food and cash crop around the world. It is considered to be an allotetraploid (2n = 4x = 40) originated from a single hybridization event between two wild diploids. The most probable hypothesis gave A. duranensis as the wild donor of the A genome and A. ipaënsis as the wild donor of the B genome. A low level of molecular polymorphism is found in cultivated germplasm and up to date few genetic linkage maps have been published. The utilization of wild germplasm in breeding programs has received little attention due to the reproductive barriers between wild and cultivated species and to the technical difficulties encountered in making large number of crosses. We report here the development of a SSR based genetic map and the analysis of genome-wide segment introgressions into the background of a cultivated variety through the utilization of a synthetic amphidiploid between A. duranensis and A. ipaënsis.Results
Two hundred ninety eight (298) loci were mapped in 21 linkage groups (LGs), spanning a total map distance of 1843.7 cM with an average distance of 6.1 cM between adjacent markers. The level of polymorphism observed between the parent of the amphidiploid and the cultivated variety is consistent with A. duranensis and A. ipaënsis being the most probable donor of the A and B genomes respectively. The synteny analysis between the A and B genomes revealed an overall good collinearity of the homeologous LGs. The comparison with the diploid and tetraploid maps shed new light on the evolutionary forces that contributed to the divergence of the A and B genome species and raised the question of the classification of the B genome species. Structural modifications such as chromosomal segment inversions and a major translocation event prior to the tetraploidisation of the cultivated species were revealed. Marker assisted selection of BC1F1 and then BC2F1 lines carrying the desirable donor segment with the best possible return to the background of the cultivated variety provided a set of lines offering an optimal distribution of the wild introgressions.Conclusion
The genetic map developed, allowed the synteny analysis of the A and B genomes, the comparison with diploid and tetraploid maps and the analysis of the introgression segments from the wild synthetic into the background of a cultivated variety. The material we have produced in this study should facilitate the development of advanced backcross and CSSL breeding populations for the improvement of cultivated peanut. 相似文献995.
The sub-cellular feruloylation and oxidative coupling sites of cell wall polysaccharides were investigated in planta by monitoring
the kinetics of appearance of arabinosyl- and feruloyl-radiolabelled polysaccharides in the protoplasmic compartment and their
secretion in the wall either in the presence or absence of brefeldin A (BFA). By using root apical segments excised from wheat
seedlings (Triticum durum Desf.), incubated with trans-[U-14C]cinnamic acid, we demonstrated that [14C]ferulate, likely [14C]diferulate, as well as trimers and larger products of ferulate are incorporated into the protoplasmic polysaccharides very
rapidly within 1–3 min of [14C]cinnamate feeding. This agrees with the assumption that (glucurono)arabinoxylans [(G)AX] feruloylation and oxidative coupling
occur intracellularly, likely in the Golgi apparatus. Simultaneously, polymer bound radioactive hydroxycinnamic acids appeared
to be incorporated into the cell wall of root apical segments as early as 2 min after trans-[U-14C]cinnamic acid feeding. On the contrary, starting from l-[1-14C]arabinose as tracer, the secretion of the pentose-containing polymers into the wall was between 5 to 10 min. These results
indicated that (G)AX feruloylation and oxidative coupling occur both intra-protoplasmically and in muro. The occurrence of
in muro feruloylation and oxidative coupling was confirmed by the use of BFA a well known inhibitor of secretion. The drug
caused a strong inhibition of the synthesis and secretion into the wall of the 14C-pentosyl-labelled polymers as well as of 14C-feruloyl-polymers. In spite of this, the total amount of 14C-feruloyl-polymers incorporated into the wall was only slightly affected by BFA. This indicates the existence of a mechanism
involved into secretion of the activated hydroxycinnamoyl precursors to the wall, alternative to that involved in polysaccharide
secretion.
Lucia Ilenia Mastrangelo and Marcello Salvatore Lenucci equally contributed to this work. 相似文献
996.
Marcello Clerici André Mourão Irina Gutsche Niels H Gehring Matthias W Hentze Andreas Kulozik Jan Kadlec Michael Sattler Stephen Cusack 《The EMBO journal》2009,28(15):2293-2306
Nonsense‐mediated decay (NMD) is a eukaryotic quality control mechanism that degrades mRNAs carrying premature stop codons. In mammalian cells, NMD is triggered when UPF2 bound to UPF3 on a downstream exon junction complex interacts with UPF1 bound to a stalled ribosome. We report structural studies on the interaction between the C‐terminal region of UPF2 and intact UPF1. Crystal structures, confirmed by EM and SAXS, show that the UPF1 CH‐domain is docked onto its helicase domain in a fixed configuration. The C‐terminal region of UPF2 is natively unfolded but binds through separated α‐helical and β‐hairpin elements to the UPF1 CH‐domain. The α‐helical region binds sixfold more weakly than the β‐hairpin, whereas the combined elements bind 80‐fold more tightly. Cellular assays show that NMD is severely affected by mutations disrupting the beta‐hairpin binding, but not by those only affecting alpha‐helix binding. We propose that the bipartite mode of UPF2 binding to UPF1 brings the ribosome and the EJC in close proximity by forming a tight complex after an initial weak encounter with either element. 相似文献
997.
Marcello Tonelli Brenda Hemmelgarn Tony Reiman Braden Manns M. Neil Reaume Anita Lloyd Natasha Wiebe Scott Klarenbach 《CMAJ》2009,180(11):E62-E71
Background
Erythropoiesis-stimulating agents are used to treat anemia in patients with cancer. However, their safety and effectiveness is controversial. We did a systematic review of the clinical efficacy and harms of these agents in adults with anemia related to cancer or chemotherapy.Methods
We conducted a systematic review of published and unpublished randomized controlled trials (RCTs) using accepted methods for literature searches, article selection, data extraction and quality assessment. We included RCTs involving anemic adults with cancer. We compared the use of erythropoiesis-stimulating agents with nonuse and assessed clinical outcomes (all-cause mortality, cardiovascular events and hypertension, health-related quality of life, blood transfusions and tumour response) and harms (serious adverse events) between groups.Results
We identified 52 trials (n = 12 006) that met our selection criteria. The pooled all-cause mortality during treatment was significantly higher in the group receiving erythropoiesis-stimulating therapy than in the control group (relative risk [RR] 1.15, 95% confidence interval [CI] 1.03 to 1.29). Compared with no treatment, use of erythropoiesis-stimulating agents led to clinically detectable improvements in disease-specific measures of quality of life. It also reduced the use of blood transfusions (RR 0.64, 95% CI 0.56 to 0.73). However, it led to an increased risk of thrombotic events (RR 1.69, 95% CI 1.27 to 2.24) and serious adverse events (RR 1.16, 95% CI 1.08 to 1.25).Interpretation
Use of erythropoiesis-stimulating agents in patients with cancer-related anemia improved some disease-specific measures of quality of life and decreased the use of blood transfusions. However, it increased the risk of death and serious adverse events. Our findings suggest that such therapy not be used routinely as an alternative to blood transfusion in patients with anemia related to cancer.Anemia related to cancer may be due to the cancer itself or it may be a complication of chemotherapy. Because anemia is associated with adverse clinical outcomes in people with cancer, including impaired quality of life1 and decreased survival,2 treatment with erythropoiesis-stimulating agents has been widely used. These agents are costly, and reimbursement policies for their use in patients with cancer-related anemia vary across Canadian jurisdictions. Recent studies suggest that their use in such patients may be associated with an increased risk of adverse events such as thromboembolism.3 Potential adverse effects have also been identified in patients with chronic kidney disease.4,5Therefore, an assessment of the efficacy and harms of erythropoiesis-stimulating agents in patients with cancer-related anemia would be useful to clinicians, and to jurisdictions that seek to develop an evidence-based reimbursement policy for these drugs. We conducted a systematic review based on work done for the Canadian Agency for Drugs and Technologies in Health6 to summarize the clinical efficacy and harms of these agents in adults with anemia related to cancer. 相似文献998.
Muscari C Grossi L Giordano E Ferrari D Bonafe F Guarnieri C Caldarera CM 《Life sciences》2003,74(1):109-123
Aim of this study was to investigate the cardiac release of nitric oxide (NO) before and after cold cardioplegia by a novel electron paramagnetic resonance (EPR) technique. Isolated rat hearts were perfused for 20 min in a Langendorff apparatus and then subjected to 3 hours potassium-hypotermic cardioplegia, followed by 20 min reperfusion. The coronary effluent was collected in a flask containing ferrous-bis-diethyldithiocarbamate as a spin trap of NO. Since the trapping agent was not delivered to the heart with the perfusion medium, we avoided that an abnormal extraction of NO from the tissue could inhibit its biological activity. The EPR signal was well detectable after equilibration (25.6 +/- 3.0 nmol/L +/- S.E.M.) and significantly increased following perfusion with 10 micromol/L serotonin (41.1 +/- 3.2 nmol/L) or 10 micromol/L nitroprusside (43.5 +/- 2.9 nmol/L). The basal level of NO did not change after reperfusion, but serotonin administration was not able to stimulate its release. Serotonin failure to stimulate NO production was not due to a loss of endothelial NO synthase, since its protein expression was not modified after reperfusion. The perfusion pressure increased by 51% after reperfusion and was quite completely restored following serotonin or nitroprusside treatment, with respect to the non-stimulated equilibration condition. Therefore, we suggest that the coronary spasm following a cold cardioplegic arrest is not due to an impaired production of basal NO and that NO-donors can be effective in relaxing vascular smooth muscle cells. 相似文献
999.
Mazzeo MF Sorrentino A Gaita M Cacace G Di Stasio M Facchiano A Comi G Malorni A Siciliano RA 《Applied and environmental microbiology》2006,72(2):1180-1189
A methodology based on matrix-assisted laser desorption ionization-time of flight mass spectrometry of intact bacterial cells was used for rapid discrimination of 24 bacterial species, and detailed analyses to identify Escherichia coli O157:H7 were carried out. Highly specific mass spectrometric profiles of pathogenic and nonpathogenic bacteria that are well-known major food contaminants were obtained, uploaded in a specific database, and made available on the Web. In order to standardize the analytical protocol, several experimental, sample preparation, and mass spectrometry parameters that can affect the reproducibility and accuracy of data were evaluated. Our results confirm the conclusion that this strategy is a powerful tool for rapid and accurate identification of bacterial species and that mass spectrometric methodologies could play an essential role in polyphasic approaches to the identification of pathogenic bacteria. 相似文献
1000.
We evaluated determinants of anti-benzo[a]pyrenediolepoxide-(B[a]PDE)-DNA adduct formation (adduct induced by the ultimate carcinogenic metabolite of B[a]P) in lymphomonocytes of subjects environmentally exposed to low doses of polycyclic aromatic hydrocarbons (PAHs) (B[a]P). Our study population consisted of 585 Caucasian subjects, all municipal workers living in North-East Italy and recruited during their periodic check-ups after informed consent. PAH (B[a]P) exposure was assessed by questionnaire. Anti-B[a]PDE-DNA levels were measured by HPLC fluorescence analysis. We found that cigarette smoking (smokers (22%) versus non-smokers, p<0.0001), dietary intake of PAH-rich meals (> or =52 (38%) versus <52 times/year, p<0.0001), and outdoor exposure (> or =4 (19%) versus <4h/day; p=0.0115) significantly influenced adduct levels. Indoor exposure significantly increased the frequency of positive subjects (> or =0.5 adducts/10(8) nucleotides; chi(2) for linear trend, p=0.051). In linear multiple regression analysis the major determinants of increased DNA adduct levels (ln values) were smoking (t=6.362, p<0.0001) and diet (t=4.035, p<0.0001). In this statistical analysis, indoor and outdoor exposure like other factors of PAH exposure had no influence. In non-smokers, the influence of diet (p<0.0001) and high indoor exposure (p=0.016) on anti-B[a]PDE-DNA adduct formation became more evident, but not that of outdoor exposure, as was confirmed by linear multiple regression analysis (diet, t=3.997, p<0.0001 and high indoor exposure, t=2.522, p=0.012). This study indicates that anti-B[a]PDE-DNA adducts can be detected in the general population and are modulated by PAH (B[a]P) exposure not only with smoking - information already known from studies with limited number of subjects - but also with dietary habits and high indoor exposure. In non-smokers, these two factors are the principal determinants of DNA adduct formation. The information provided here seems to be important, since DNA adduct formation in surrogate tissue is an index of genotoxic exposure also in target organs (e.g., lung) and their increase may also be predictive of higher risk for PAH-related cancers. 相似文献