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101.
Liu Z Ramanoudjame G Liu D Fox RO Jayaraman V Kurnikova M Cascio M 《Biochemistry》2008,47(37):9803-9810
A novel truncated form (residues 1-214, with a randomized C-terminal tail) of the ligand-binding extracellular domain (ECD) of the human alpha1 glycine receptor (GlyR), with amino acids from the corresponding sequence of an acetylcholine binding protein (AChBP) substituted for two relatively hydrophobic membrane-proximal loops, was overexpressed using a baculovirus expression system. The mutant GlyR ECD, named GlyBP, was present in both soluble and membrane-associated fractions after cell lysis, though only the latter appeared to be in a native-like conformation capable of binding strychnine, a GlyR specific antagonist. The membrane-associated GlyBP was solubilized, and detergent/lipid/protein micelles were affinity purified. After detergent removal, GlyBP may be isolated in either aqueous or vesicular form. Binding assays and spectroscopic studies using circular dichroism and FRET are consistent with both forms adopting equivalent native-like conformations. Thus, GlyBP may be isolated as a soluble or membrane-associated assembly that serves as a structural and functional homologue of the ECD of GlyR. 相似文献
102.
D R Hickey K Jayaraman C T Goodhue J Shah S A Fingar J M Clements Y Hosokawa S Tsunasawa F Sherman 《Gene》1991,105(1):73-81
Genes encoding tuna, pigeon, and horse cytochromes c were constructed with synthetic oligodeoxyribonucleotides having preferred codons and portions of the iso-1-cytochrome c-encoding gene from the yeast Saccharomyces cerevisiae. The genes were ligated into an expression vector, which contains the normal 5'- and 3'-untranslated regions of the yeast iso-1-cytochrome c gene, and were integrated in single copy into the chromosome. Yeast strains were also constructed with multiple integrated copies of the pigeon gene. The heterologous and normal mRNA levels of the single-copy strains were equivalent. Although the N-terminal methionines were completely cleaved in the heterospecific proteins, the levels of trimethylation of Lys72 and acetylation of N-terminal glycines ranged from 39-78% and 10-70%, respectively. Horse cytochrome c was produced at a nearly normal level, whereas the pigeon and tuna cytochromes c were produced at approx. 40% of the normal levels. The levels of the cytochromes c and growth of the mutant yeast strains indicated that the heterospecific cytochromes c had approx. 50% specific activity in vivo. 相似文献
103.
104.
105.
106.
V K Jayaraman 《Biotechnology progress》1992,8(5):462-464
A methodology for simplifying the solution procedure for hollow fiber bioreactor design equations has been described. Such a procedure facilitates decoupling of membrane and spongy matrix equations from the tube side equations. The equivalence between the reduced equations and the hemodialyzer problem has been explicitly obtained. 相似文献
107.
108.
Drew M. Dolino Swarna S. Ramaswamy Vasanthi Jayaraman 《Journal of visualized experiments : JoVE》2014,(91)
Luminescence Resonance Energy Transfer, or LRET, is a powerful technique used to measure distances between two sites in proteins within the distance range of 10-100 Å. By measuring the distances under various ligated conditions, conformational changes of the protein can be easily assessed. With LRET, a lanthanide, most often chelated terbium, is used as the donor fluorophore, affording advantages such as a longer donor-only emission lifetime, the flexibility to use multiple acceptor fluorophores, and the opportunity to detect sensitized acceptor emission as an easy way to measure energy transfer without the risk of also detecting donor-only signal. Here, we describe a method to use LRET on membrane proteins expressed and assayed on the surface of intact mammalian cells. We introduce a protease cleavage site between the LRET fluorophore pair. After obtaining the original LRET signal, cleavage at that site removes the specific LRET signal from the protein of interest allowing us to quantitatively subtract the background signal that remains after cleavage. This method allows for more physiologically relevant measurements to be made without the need for purification of protein. 相似文献
109.
Hierarchical Zn–Co–S Nanowires as Advanced Electrodes for All Solid State Asymmetric Supercapacitors 下载免费PDF全文
A facile two‐step strategy is developed to design the large‐scale synthesis of hierarchical, unique porous architecture of ternary metal hydroxide nanowires grown on porous 3D Ni foam and subsequent effective sulfurization. The hierarchical Zn–Co–S nanowires (NWs) arrays are directly employed as an electrode for supercapacitors application. The as‐synthesized Zn–Co–S NWs deliver an ultrahigh areal capacity of 0.9 mA h cm?2 (specific capacity of 366.7 mA h g?1) at a current density of 3 mA cm?2, with an exceptional rate capability (≈227.6 mA h g?1 at a very high current density of 40 mA cm?2) and outstanding cycling stability (≈93.2% of capacity retention after 10 000 cycles). Most significantly, the assembled Zn–Co–S NWs//Fe2O3@reduced graphene oxide asymmetric supercapacitors with a wide operating potential window of ≈1.6 V yield an ultrahigh volumetric capacity of ≈1.98 mA h cm?3 at a current density of 3 mA cm?2, excellent energy density of ≈81.6 W h kg?1 at a power density of ≈559.2 W kg?1, and exceptional cycling performance (≈92.1% of capacity retention after 10 000 cycles). This general strategy provides an alternative to design the other ternary metal sulfides, making it facile, free‐standing, binder‐free, and cost‐effective ternary metal sulfide‐based electrodes for large‐scale applications in modern electronics. 相似文献
110.
Achary A Hariharan KA Bandhyopadhyaya S Ramachandran R Jayaraman K 《Biotechnology and bioengineering》1997,55(1):148-154
D-Hydantoinases (E.C.3.5.2.2) are commercially valuable enzymes involved in the production of D-amino acids. However, commercial exploitation of the biological process is rare, mainly because sufficient details are not available on the efficient production of these enzymes by microorganisms. In the present study, Agrobacterium radiobacter was used as the source of D-hydantoinase and its production was optimized with inexpensive carbon and nitrogen sources. The four media components selected to study their effect on biomass and/or enzyme activities were molasses, ammonium nitrate, sodium di-hydrogen orthophosphate, and manganese chloride. With the use of an empirical modeling technique (response surface method), we have optimized both biomass and enzyme production in this organism, with a minimal number of batches. Experiments were performed with optimized media components to validate the model. The maximum level of enzyme and biomass obtained was 35 U/mL and 1.69 mg/mL, respectively. (c) 1997 John Wiley & Sons, Inc. Biotechnol Bioeng 55: 148-154, 1997. 相似文献