全文获取类型
收费全文 | 4921篇 |
免费 | 301篇 |
国内免费 | 2篇 |
专业分类
5224篇 |
出版年
2024年 | 8篇 |
2023年 | 16篇 |
2022年 | 71篇 |
2021年 | 80篇 |
2020年 | 52篇 |
2019年 | 71篇 |
2018年 | 100篇 |
2017年 | 86篇 |
2016年 | 156篇 |
2015年 | 263篇 |
2014年 | 318篇 |
2013年 | 334篇 |
2012年 | 484篇 |
2011年 | 424篇 |
2010年 | 269篇 |
2009年 | 240篇 |
2008年 | 336篇 |
2007年 | 263篇 |
2006年 | 257篇 |
2005年 | 220篇 |
2004年 | 190篇 |
2003年 | 169篇 |
2002年 | 148篇 |
2001年 | 127篇 |
2000年 | 124篇 |
1999年 | 112篇 |
1998年 | 39篇 |
1997年 | 28篇 |
1996年 | 20篇 |
1995年 | 21篇 |
1994年 | 20篇 |
1993年 | 17篇 |
1992年 | 26篇 |
1991年 | 23篇 |
1990年 | 14篇 |
1989年 | 11篇 |
1988年 | 8篇 |
1987年 | 4篇 |
1986年 | 9篇 |
1984年 | 4篇 |
1982年 | 4篇 |
1981年 | 3篇 |
1977年 | 4篇 |
1976年 | 4篇 |
1974年 | 7篇 |
1973年 | 3篇 |
1972年 | 3篇 |
1969年 | 4篇 |
1967年 | 5篇 |
1966年 | 5篇 |
排序方式: 共有5224条查询结果,搜索用时 0 毫秒
141.
Traditionally, the population of human embryonic stem cell (hESC) culture is estimated through haemacytometer counts, which include harvesting the cells and manually analyzing a fraction of an entire population. Obviously, through this highly invasive method, it is not possible to preserve any spatial information on the cell population. The goal of this study is to identify a fast and consistent method for in situ automated hESC population estimation to quantitatively estimate the cell growth. Therefore, cell cultures were fixed, stained, and their nuclei imaged through high‐resolution microscopy, and the images were processed with different image analysis techniques. The proposed method first identifies signal and background by computing an image specific threshold for image segmentation. By applying a morphological operator (watershed), we split most physically overlapping nuclei, leading to a pixel area distribution of isolated signal areas on the image. On the basis of this distribution, we derive a nucleus area model, describing the distribution of the area of cell debris, single nuclei, and small groups of connected nuclei. Through the model, we can give a quantitative estimation of the population. The focus of this study is on low‐density human embryonic stem cell populations; hence cultures were measured at days 2–3 after seeding. Compared with manual cell counts, the automatic method achieved higher accuracy with <6% error. © 2009 American Institute of Chemical Engineers Biotechnol. Prog., 2010 相似文献
142.
Y.‐R. Lim R.‐Y. Yoon E.‐S. Seo Y.‐S. Kim C.‐S. Park D.‐K. Oh 《Journal of applied microbiology》2010,109(4):1188-1197
Aims: To characterize of a thermostable recombinant α‐l ‐arabinofuranosidase from Caldicellulosiruptor saccharolyticus for the hydrolysis of arabino‐oligosaccharides to l ‐arabinose. Methods and Results: A recombinant α‐l ‐arabinofuranosidase from C. saccharolyticus was purified by heat treatment and Hi‐Trap anion exchange chromatography with a specific activity of 28·2 U mg?1. The native enzyme was a 58‐kDa octamer with a molecular mass of 460 kDa, as measured by gel filtration. The catalytic residues and consensus sequences of the glycoside hydrolase 51 family of α‐l ‐arabinofuranosidases were completely conserved in α‐l ‐arabinofuranosidase from C. saccharolyticus. The maximum enzyme activity was observed at pH 5·5 and 80°C with a half‐life of 49 h at 75°C. Among aryl‐glycoside substrates, the enzyme displayed activity only for p‐nitrophenyl‐α‐l ‐arabinofuranoside [maximum kcat/Km of 220 m(mol l?1)?1 s?1] and p‐nitrophenyl‐α‐l ‐arabinopyranoside. This substrate specificity differs from those of other α‐l ‐arabinofuranosidases. In a 1 mmol l?1 solution of each sugar, arabino‐oligosaccharides with 2–5 monomer units were completely hydrolysed to l ‐arabinose within 13 h in the presence of 30 U ml?1 of enzyme at 75°C. Conclusions: The novel substrate specificity and hydrolytic properties for arabino‐oligosaccharides of α‐l ‐arabinofuranosidase from C. saccharolyticus demonstrate the potential in the commercial production of l ‐arabinose in concert with endoarabinanase and/or xylanase. Significance and Impact of the Study: The findings of this work contribute to the knowledge of hydrolytic properties for arabino‐oligosaccharides performed by thermostable α‐l ‐arabinofuranosidase. 相似文献
143.
The recombinant β-carotene 15,15′-monooxygenase from chicken liver was purified as a single 60 kDa band by His-Trap HP and Resource Q chromatography.
It had a molecular mass of 240 kDa by gel filtration indicating the native form to be tetramer. The enzyme converted β-carotene under maximal conditions (pH 8.0 and 37°C) with a k
cat of 1.65 min−1 and a K
m of 26 μM and its conversion yield of β-carotene to retinal was 120% (mol mol−1). The enzyme displayed catalytic efficiency and conversion yield for β-carotene, β-cryptoxanthin, β-apo-8′-carotenal, β-apo-4′-carotenal, α-carotene and γ-carotene in decreasing order but not for zeaxanthin, lutein, β-apo-12′-carotenal and lycopene, suggesting that the presence of one unsubstituted β-ionone ring in a substrate with a molecular weight greater than C30 seems to be essential for enzyme activity. 相似文献
144.
Phosphorylation of focal adhesion kinase at tyrosine 861 is crucial for Ras transformation of fibroblasts 总被引:4,自引:0,他引:4
Lim Y Han I Jeon J Park H Bahk YY Oh ES 《The Journal of biological chemistry》2004,279(28):29060-29065
Although elevated expression and increased tyrosine phosphorylation of focal adhesion kinase (FAK) are crucial for tumor progression, the mechanism by which FAK promotes oncogenic transformation is unclear. We have therefore determined the role of FAK phosphorylation at tyrosine 861 in the oncogenic transformation of NIH3T3 fibroblasts. FAK phosphorylation at tyrosine 861 was increased in both constitutively H-Ras-transformed and H-Ras-inducible NIH3T3 cells, in parallel with cell transformation. However, H-Ras-inducible cells transfected with the nonphosphorylatable mutant FAK Y861F showed decreased migration/invasion, focus forming activity and anchorage-independent growth, compared with either wild-type or kinase-defective FAK. In contrast to unaltered FAK/Src activity, the association of FAK and p130(CAS) was decreased in FAK Y861F-transfected cells, and FAK phosphorylation at tyrosine 861 enhanced this association in vitro. Consistently, FAK Y861F-transfected cells were defective in activation of c-Jun NH(2)-terminal kinase and in expression of matrix metalloproteinase-9 during transformation. Taken together, these results strongly suggest that FAK phosphorylation at tyrosine 861 is crucial for H-Ras-induced transformation through regulation of the association of FAK with p130(CAS). 相似文献
145.
146.
The rpiB gene, encoding ribose-5-phosphate isomerase (RpiB) from Clostridium thermocellum, was cloned and expressed in Escherichia coli. RpiB converted d-psicose into d-allose but it did not convert d-xylose, l-rhamnose, d-altrose or d-galactose. The production of d-allose by RpiB was maximal at pH 7.5 and 65°C for 30 min. The half-lives of the enzyme at 50°C and 65°C were 96 h and 4.7 h,
respectively. Under stable conditions of pH 7.5 and 50°C, 165 g d-allose l−1
was produced without by-products from 500 g d-psicose l−1 after 6 h. 相似文献
147.
148.
149.
Oh M Im I Lee YJ Kim YH Yoon JH Park HG Higashiyama S Kim YC Park WJ 《Bioorganic & medicinal chemistry letters》2004,14(24):6071-6074
We describe a series of potent and selective inhibitors of ADAM12 that were discovered using computational screening of a focused virtual library. The initial structure-based virtual screening selected 64 compounds from a 3D database of 67,062 molecules. Being evaluated by a cell-based ADAM12 activity assay, compounds 5, 11, 14, 16 were further identified as the potent and selective inhibitors of ADAM12 with low nanomolar IC50 values. The mechanism underlying the potency and selectivity of a representative compound, 5, was investigated through molecular docking studies. 相似文献
150.
In this study, the cellular responses of Stenotrophomonas sp. OK-5 to explosive 2,4,6-trinitrotoluene (TNT) have been extensively analyzed. The stress shock proteins, which might contribute to enhancing cellular resistance to TNT-mediated toxicity, were induced at different concentrations of TNT used as a substrate for cell culture of Stenotrophomonas sp. OK-5 capable of utilizing TNT. Proteomic analysis for 2-DE of soluble protein fractions from the culture of OK-5 exposed to TNT demonstrated approximately 300 spots on the silver-stained gel ranging from pH 3 to pH 10. Among them, 10 spots significantly induced and expressed in response to TNT were selected and analyzed. As the result of internal amino acid sequencing with ESI-Q TOF mass spectrometry, TNT-mediated stress shock proteins such as DnaK, OmpW, and OsmC were identified and characterized. Survival of strain OK-5 was periodically monitored in the presence of different concentrations of TNT along with the production of the stress shock proteins. Cells of strain OK-5 pre-exposed to TNT had in improved survival tolerance. Analysis of total cellular fatty acids in strain OK-5 suggested that several saturated or unsaturated fatty acids might increase or decrease under TNT-mediated stress condition. Scanning electron microscopy of cells treated with 0.8 mM TNT for 12 h revealed irregular rod shapes with wrinkled surfaces. 相似文献