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111.
Parallel Characterization of Anaerobic Toluene- and Ethylbenzene-Degrading Microbial Consortia by PCR-Denaturing Gradient Gel Electrophoresis, RNA-DNA Membrane Hybridization, and DNA Microarray Technology 总被引:2,自引:9,他引:2 下载免费PDF全文
Yoshikazu Koizumi John J. Kelly Tatsunori Nakagawa Hidetoshi Urakawa Saïd El-Fantroussi Saleh Al-Muzaini Manabu Fukui Yoshikuni Urushigawa David A. Stahl 《Applied microbiology》2002,68(7):3215-3225
A mesophilic toluene-degrading consortium (TDC) and an ethylbenzene-degrading consortium (EDC) were established under sulfate-reducing conditions. These consortia were first characterized by denaturing gradient gel electrophoresis (DGGE) fingerprinting of PCR-amplified 16S rRNA gene fragments, followed by sequencing. The sequences of the major bands (T-1 and E-2) belonging to TDC and EDC, respectively, were affiliated with the family Desulfobacteriaceae. Another major band from EDC (E-1) was related to an uncultured non-sulfate-reducing soil bacterium. Oligonucleotide probes specific for the 16S rRNAs of target organisms corresponding to T-1, E-1, and E-2 were designed, and hybridization conditions were optimized for two analytical formats, membrane and DNA microarray hybridization. Both formats were used to characterize the TDC and EDC, and the results of both were consistent with DGGE analysis. In order to assess the utility of the microarray format for analysis of environmental samples, oil-contaminated sediments from the coast of Kuwait were analyzed. The DNA microarray successfully detected bacterial nucleic acids from these samples, but probes targeting specific groups of sulfate-reducing bacteria did not give positive signals. The results of this study demonstrate the limitations and the potential utility of DNA microarrays for microbial community analysis. 相似文献
112.
Globular proteins have high packing densities as a result of residue side chains in the core achieving a tight, complementary packing. The internal packing is considered the main determinant of native protein structure. From that point of view, we present here a method of energy minimization using an automata network to predict a set of amino acid sequences and their side-chain conformations from a desired backbone geometry for de novo design of proteins. Using discrete side-chain conformations, that is, rotamers, the sequence generation problem from a given backbone geometry becomes one of combinatorial problems. We focused on the residues composing the interior core region and predicted a set of amino acid Sequences and their side-chain conformations only from a given backbone geometry. The kinds of residues were restricted to six hydrophobic amino acids (Ala, Ile, Met, Leu, Phe, and Val) because the core regions are almost always composed of hydrophobic residues. The obtained sequences were well packed as was the native sequence. The method can be used for automated sequence generation in the de novo design of proteins. © 1994 Wiley-Liss, Inc. 相似文献
113.
114.
At least two polymorphic Alu insertions have been previously identified and characterized within the class I region of the
major histocompatibility complex (MHC). We have identified another two new polymorphic Alu insertions, AluyHJ and AluyHF,
located near HLA-J and HLA-F, respectively, within the a block of the MHC. Here we report on (1) the haplotypic relationships
between the Alu dimorphisms and the HLA-A locus within a panel of 51 IHW homozygous cell lines representing at least 36 HLA
class I haplotypes, (2) the Alu genotype, allele, and haplotype frequencies present in the Australian Caucasians and Japanese
populations, and (3) the frequency of association between the different Alu dimorphisms and the HLA-A alleles in 109 Australian
Caucasians and 99 Japanese. PCR was used to detect the presence or absence of insertion for AluyHJ, AluyHG, and AluyHF within
the DNA samples prepared from the cell lines and the two population groups that had been previously typed for HLA-A. In the
homozygous cell lines, all three Alu insertions were found in only one HLA class I haplotype (HLA-A1, -B57, -Cw6), no Alu
insertions were detected in six HLA class I haplotypes and one or more of the Alu insertions were found in 29 HLA class I
haplotypes. At least one of the Alu insertions was found in about 86% of the Japanese and Australian individuals, with the
AluyHJ generally related inversely to AluyHG and/or AluyHF. The gene frequency of the AluyHJ and AluyHF insertions was significantly
different (p <0.05) BETWEEN JAPANESE AND AUSTRALIANS, WHEREAS THERE WAS NO DIFFERENCE (P > 0.05) between the frequencies of
AluyHG in the two populations. The Alu haplotype frequencies were also significantly different between the Japanese and the
Australians. In the cell lines and the population groups, the AluyHJ insertion was most frequently found associated with HLA-A1
or A24, AluyHG with HLA-A2, and AluyHF with HLA-A2, -A10, or -A26. This study suggests that the three polymorphic Alu elements
have been inserted into the a block of the MHC in different progenitor groups and therefore will be useful lineage and linkage
markers in human population studies and for elucidating the evolution of HLA class I haplotypes. 相似文献
115.
Ammonia-oxidizing bacteria on root biofilms and their possible contribution to N use efficiency of different rice cultivars 总被引:12,自引:0,他引:12
Briones Aurelio M. Okabe Satoshi Umemiya Yoshiaki Ramsing Niels-Birger Reichardt Wolfgang Okuyama Hidetoshi 《Plant and Soil》2003,250(2):335-348
Ammonia-oxidizing bacteria (AOB) populations were studied on the root surface of different rice cultivars by PCR coupled with denaturing gradient gel electrophoresis (DGGE) and fluorescence in situ hybridization (FISH). PCR-DGGE of the ammonium monooxygenase gene (amoA) showed a generally greater diversity on root samples compared to rhizosphere and unplanted soil. Sequences affiliated with Nitrosomonas spp. tended to be associated with modern rice hybrid lines. Root-associated AOB observed by FISH were found within a discrete biofilm coating the root surface. Although the total abundance of AOB on root biofilms of different rice cultivars did not differ significantly, there were marked contrasts in their population structure, indicating selection of Nitrosomonas spp. on roots of a hybrid cultivar. Observations by FISH on the total bacterial community also suggested that different rice cultivars support different bacterial populations even under identical environmental conditions. The presence of active AOB in the root environment predicts that a significant proportion of the N taken up by certain rice cultivars is in the form of NO3
–-N produced by the AOB. Measurement of plant growth of hydroponically grown plants showed a stronger response of hybrid cultivars to the co-provision of NH4
+ and NO3
–. In soil-grown plants, N use efficiency in the hybrid was improved during ammonium fertilization compared to nitrate fertilization. Since ammonium-fertilized plants actually receive a mixture of NH4
+ and NO3
– with ratios depending on root-associated nitrification activity, these results support the advantage of co-provision of ammonium and nitrate for the hybrid cultivar. 相似文献
116.
Masamitsu Konno Atsushi Hamabe Shinichiro Hasegawa Hisataka Ogawa Takahito Fukusumi Shimpei Nishikawa Katsuya Ohta Yoshihiro Kano Miyuki Ozaki Yuko Noguchi Daisuke Sakai Toshihiro Kudoh Koichi Kawamoto Hidetoshi Eguchi Taroh Satoh Masahiro Tanemura Hiroaki Nagano Yuichiro Doki Masaki Mori Hideshi Ishii 《Development, growth & differentiation》2013,55(3):309-318
Adipose tissue‐derived mesenchymal stem cells (ADSCs) are multipotent and can differentiate into various cell types, including osteocytes, adipocytes, neural cells, vascular endothelial cells, cardiomyocytes, pancreatic β‐cells, and hepatocytes. Compared with the extraction of other stem cells such as bone marrow‐derived mesenchymal stem cells (BMSCs), that of ADSCs requires minimally invasive techniques. In the field of regenerative medicine, the use of autologous cells is preferable to embryonic stem cells or induced pluripotent stem cells. Therefore, ADSCs are a useful resource for drug screening and regenerative medicine. Here we present the methods and mechanisms underlying the induction of multilineage cells from ADSCs. 相似文献
117.
Kohei Yuyama Hui Sun Shota Sakai Susumu Mitsutake Megumi Okada Hidetoshi Tahara Jun-ichi Furukawa Naoki Fujitani Yasuro Shinohara Yasuyuki Igarashi 《The Journal of biological chemistry》2014,289(35):24488-24498
Elevated levels of amyloid-β peptide (Aβ) in the human brain are linked to the pathogenesis of Alzheimer disease. Recent in vitro studies have demonstrated that extracellular Aβ can bind to exosomes, which are cell-secreted nanovesicles with lipid membranes that are known to transport their cargos intercellularly. Such findings suggest that the exosomes are involved in Aβ metabolism in brain. Here, we found that neuroblastoma-derived exosomes exogenously injected into mouse brains trapped Aβ and with the associated Aβ were internalized into brain-resident phagocyte microglia. Accordingly, continuous intracerebral administration of the exosomes into amyloid-β precursor protein transgenic mice resulted in marked reductions in Aβ levels, amyloid depositions, and Aβ-mediated synaptotoxicity in the hippocampus. In addition, we determined that glycosphingolipids (GSLs), a group of membrane glycolipids, are highly abundant in the exosomes, and the enriched glycans of the GSLs are essential for Aβ binding and assembly on the exosomes both in vitro and in vivo. Our data demonstrate that intracerebrally administered exosomes can act as potent scavengers for Aβ by carrying it on the exosome surface GSLs and suggest a role of exosomes in Aβ clearance in the central nervous system. Improving Aβ clearance by exosome administration would provide a novel therapeutic intervention for Alzheimer disease. 相似文献
118.
119.
Inoue H Iihara A Takahashi H Shimada I Ishida I Maeda Y 《Protein science : a publication of the Protein Society》2011,20(12):1971-1981
VHH is the binding domain of the IgG heavy chain. Some VHHs have an extremely long CDR3 that contributes to antigen binding. We studied the antigen binding ability of CDR3 by grafting a CDR3 from an antigen-binding VHH onto a nonbinding VHH. cAb-CA05-(1RI8), the CDR3-grafted VHH, had an antigen-binding ability. To find a human scaffold protein acceptable for VHH CDR3 grafting, we focused on the conserved structure of VHH, especially the N-terminal and C-terminal amino acid residues of the CDR3 loop and the Cys residue of CDR1. Human origin protein structures with the same orientation were searched in PDB and ubiquitin was selected. Ubi-(1RI8), the CDR3-grafted ubiquitin, had antigen-binding ability, though the affinity was relatively low compared to cAb-CA05-(1RI8). The thermodynamic parameters of Ubi-(1RI8) binding to HEWL were different from cAb-CA05-(1RI8). Hydrogen-deuterium exchange experiments showed decreased stability around the CDR3 grafting region of Ubi-(1RI8), which might explain the decreased antigen-binding ability and the differences in thermodynamic properties. We concluded that the orientation of the CDR3 sequence of Ubi-(1RI8) could not be reconstructed correctly. 相似文献
120.
Miura T Fukami TA Hasegawa K Ono N Suda A Shindo H Yoon DO Kim SJ Na YJ Aoki Y Shimma N Tsukuda T Shiratori Y 《Bioorganic & medicinal chemistry letters》2011,21(19):5778-5783
Heat shock protein 90 (Hsp90) is a molecular chaperone which regulates maturation and stabilization of its substrate proteins, known as client proteins. Many client proteins of Hsp90 are involved in tumor progression and survival and therefore Hsp90 can be a good target for developing anticancer drugs. With the aim of efficiently identifying a new class of orally available inhibitors of the ATP binding site of this protein, we conducted fragment screening and virtual screening in parallel against Hsp90. This approach quickly identified 2-aminotriazine and 2-aminopyrimidine derivatives as specific ligands to Hsp90 with high ligand efficiency. In silico evaluation of the 3D X-ray Hsp90 complex structures of the identified hits allowed us to promptly design CH5015765, which showed high affinity for Hsp90 and antitumor activity in human cancer xenograft mouse models. 相似文献