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241.
A pseudoknot-like structure required for efficient self-cleavage of hepatitis delta virus RNA 总被引:41,自引:0,他引:41
Hepatitis delta virus genomic and antigenomic RNAs contain a self-cleavage site hypothesized to function in processing the viral RNA during replication. Self-cleavage requires only a divalent cation and is mediated at the genomic site by a sequence of less than 85 nucleotides. We propose that the genomic self-cleaving sequence element and a corresponding sequence from the anti-genomic RNA could generate related secondary structures. The region of the antigenomic sequence, predicted from the proposed structure, was synthesized and shown to be sufficient for self-cleavage. Evidence for two stems which form a tertiary interaction was obtained by site-specific mutagenesis of the antigenomic sequence. Efficient self-cleavage in 10 M formamide or 5 M urea, also a property of the genomic sequence, was dependent on base-pairing in both stems. But in the absence of denaturants, the stem distal to the site of cleavage was not required, suggesting that the tertiary interaction stabilizes the structure required for self-cleavage. 相似文献
242.
A novel cyclin encoded by a bcl1-linked candidate oncogene 总被引:145,自引:0,他引:145
T Motokura T Bloom H G Kim H Jüppner J V Ruderman H M Kronenberg A Arnold 《Nature》1991,350(6318):512-515
We have previously identified a candidate oncogene (PRAD1 or D11S287E) on chromosome 11q13 which is clonally rearranged with the parathyroid hormone locus in a subset of benign parathyroid tumours. We now report that a cloned human placental PRAD1 complementary DNA encodes a protein of 295 amino acids with sequence similarities to the cyclins. Cyclins can form a complex with and activate p34cdc2 protein kinase, thereby regulating progress through the cell cycle. PRAD 1 messenger RNA levels vary dramatically across the cell cycle in HeLa cells. Addition of the PRAD1 protein to interphase clam embryo lysates containing inactive p34cdc2 kinase and lacking endogenous cyclins allows it to be isolated using beads bearing p13suc1, a yeast protein that binds cdc2 and related kinases with high affinity and coprecipitates kinase-associated proteins. Addition of PRAD1 also induces phosphorylation of histone H1, a preferred substrate of cdc2. These data suggest that PRAD1 encodes a novel cyclin whose overexpression may play an important part in the development of various tumours with abnormalities in 11q13. 相似文献
243.
Mutational analysis of a protein-folding pathway 总被引:6,自引:0,他引:6
The effects of amino-acid replacements on the disulphide-coupled folding pathway of bovine pancreatic trypsin inhibitor have been examined. Replacements at three sites destabilize the native protein relative to the unfolded state, but have different effects on the relative stabilities of the disulphide-bonded folding intermediates, thus allowing the roles of the altered residues during folding to be distinguished. 相似文献
244.
Inhibition of endocytic vesicle fusion in vitro by the cell-cycle control protein kinase cdc2 总被引:3,自引:0,他引:3
Membrane transport between the endoplasmic reticulum and the plasma membrane, which involves the budding and fusion of carrier vesicles, is inhibited during mitosis in animal cells. At the same time, the Golgi complex and the nuclear envelope, as well as the endoplasmic reticulum in some cell types, become fragmented. Fragmentation of the Golgi is believed to facilitate its equal partitioning between daughter cells. In fact, it has been postulated that both the inhibition of membrane traffic and Golgi fragmentation during mitosis are due to an inhibition of vesicle fusion, while vesicle budding continues. Although less is known about the endocytic pathway, internalization and receptor recycling are also arrested during mitosis. We have now used a cell-free assay to show that the fusion of endocytic vesicles from baby hamster kidney cells is reduced in Xenopus mitotic cytosol when compared with interphase cytosol. We reconstituted this inhibition in interphase cytosol by adding a preparation enriched in the starfish homologue of the cdc2 protein kinase. Inhibition was greater than or equal to 90% when the added cdc2 activity was in the range estimated for that in mitotic Xenopus eggs, which indicates that during mitosis the cdc2 kinase mediates an inhibition of endocytic vesicle fusion, and possibly other fusion events in membrane traffic. 相似文献
245.
A recombinant immunotoxin consisting of two antibody variable domains fused to Pseudomonas exotoxin 总被引:23,自引:0,他引:23
V K Chaudhary C Queen R P Junghans T A Waldmann D J FitzGerald I Pastan 《Nature》1989,339(6223):394-397
Antibodies and growth factors have been chemically coupled to different toxins to produce cytotoxic molecules that selectively kill cells bearing appropriate antigens or receptors. Antibody-toxin conjugates (immunotoxins) produced using conventional chemical coupling techniques have several undesirable characteristics. The smallest binding unit of an antibody is an Fv fragment which consists of a light and heavy chain variable domain. Recently, active single chain Fv fragments of antibodies have been produced in Escherichia coli by attaching the light and heavy chain variable domains together with a peptide linker. Here we describe the construction and expression in E. coli of a single chain antibody toxin fusion protein, anti-Tac(Fv)-PE40, in which the variable regions of anti-Tac, a monoclonal antibody to the p55 subunit of the human interleukin-2 receptor, are joined in peptide linkage to PE40, a modified form of Pseudomonas exotoxin lacking its binding domain. Anti-Tac(Fv)-PE40 was very cytotoxic to two interleukin-2 receptor-bearing human cell lines but was not cytotoxic to receptor-negative cells. 相似文献
246.
Mycoplasma infection of cultured cells 总被引:11,自引:0,他引:11
Mycoplasma contamination is tough to detect and even more difficult to eradicate. It is best to start over fresh from clean cell stocks, but several elimination options are available. 相似文献
247.
Functional cloning of ICAM-2, a cell adhesion ligand for LFA-1 homologous to ICAM-1 总被引:84,自引:0,他引:84
The leukocyte adhesion molecule LFA-1 mediates a wide range of lymphocyte, monocyte, natural killer cell, and granulocyte interactions with other cells in immunity and inflammation. LFA-1 (CD11a/CD18) is a receptor for intercellular adhesion molecule 1 (ICAM-1, CD54), a surface molecule which is constitutively expressed on some tissues and induced on other in inflammation. Induction of ICAM-1 on epithelial cells, endothelial cells and fibroblasts mediates LFA-1-dependent adhesion of lymphocytes. Several lines of evidence have suggested the existence of a second LFA-1 ligand: homotypic adhesion of one cell line was inhibited by a monoclonal antibody to LFA-1, but not by one to ICAM-1; there exists an LFA-1-dependent, ICAM-1-independent pathway of adhesion to endothelial cells; and also, there are some types of target cells in which LFA-1-dependent T-lymphocyte adhesion and lysis are independent of ICAM-1. We have cloned this second ligand, designated ICAM-2, using a novel method for identifying ligands of adhesion molecules. ICAM-2 is an integral membrane protein with two immunoglobulin-like domains, whereas ICAM-1 has five. Remarkably, ICAM-2 is much more closely related to the two most N-terminal domains of ICAM-1 (34% identity) than either ICAM-1 or ICAM-2 is to other members of the immunoglobulin superfamily, demonstrating the existence of a subfamily of immunoglobulin-like ligands that bind the same integrin receptor. 相似文献
248.
给出一种表达加权有向图的数据结构,它使得对此有向图进行“插入”操作后,只需进行O(n2)时间的维护工作,就可使得每对结点间的最短路径迅速地得到修整。 相似文献
249.
王耀青 《华中科技大学学报(自然科学版)》1994,(3)
研究一类线性多变量系统的鲁棒极点配置问题,提出了一种基于极点配置和目标函数优化的鲁棒极点配置方法。基本思想是,利用极点配置的参数化表示结果,以系统输出的L∞范数作为极点配置寻优的准则函数,通过解优化问题的途径确定极点配置问题中的自由参数。 相似文献
250.
运用气相k-ε模型和颗粒相代数方程模型对正交射流煤粉燃烧器气固两相流场进行了数值模拟,获得了该燃烧器内的气、固两相速度分布,颗粒质量流分布,并与实验值进行了对比,两者定性符合,同时,探讨了颗粒直径对颗粒混合的影响,得到了一些很有价值的结论。 相似文献