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131.
132.
HhaI and HpaII DNA methyltransferases bind DNA mismatches, methylate uracil and block DNA repair. 总被引:10,自引:7,他引:3 下载免费PDF全文
The hydrolytic deamination of 5-methylcytosine (5-mC) to thymine (T) is believed to be responsible for the high mutability of the CpG dinucleotide in DNA. We have shown a possible alternate mechanism for mutagenesis at CpG in which HpaII DNA-(cytosine-5) methyltransferase (M.HpaII) can enzymatically deaminate cytosine (C) to uracil (U) in DNA [Shen, J.-C., Rideout, W.M., III and Jones, P.A., Cell, 71, 1073-1080, (1992)]. Both the hydrolytic deamination of 5-mC and enzymatic deamination of C create premutagenic DNA mismatches (G:U and G:T) with the guanine (G) originally paired to the normal C. Surprisingly, we found that DNA-(cytosine-5) methyltransferases have higher affinities for these DNA mismatches than for their normal G:C targets and are capable of transferring a methyl group to the 5-position of U, creating T at low efficiencies. This binding by methyltransferase to mismatches at the recognition site prevented repair of G:U mismatches by uracil DNA glycosylase in vitro. 相似文献
133.
Stabilization of double-stranded oligonucleotides using backbone-linked disulfide bridges. 总被引:1,自引:1,他引:0 下载免费PDF全文
A convenient, practical route to the synthesis of disulfide-bridged oligonucleotides has been developed. Aliphatic linkers with terminal thiol groups have been attached to the phosphodiester backbones of partially or fully complementary oligonucleotide sequences and oxidized to yield covalently closed oligonucleotides with disulfide bridges. This procedure has been used to prepare a duplex with disulfide bridges at both ends and stem-loop sequences with single disulfide bridges. Oxidation of a self-complementary duplex possessing terminal thiol groups produced both hairpin and duplex structures with disulfide bridges, the relative proportions of each being dependent upon the reaction conditions. These bridged hairpin and duplex structures were shown to be interconvertible by reduction and re-oxidation. The melting profiles of disulfide-bridged oligonucleotides were compared with the same sequences without bridges and with sequences possessing triethylene glycol bridges, and in all cases the introduction of disulfide bridges resulted in a considerable increase in thermal stability. EcoRI endonuclease was capable of cleaving a disulfide-bridged duplex possessing a recognition site for this enzyme, thus supporting a lack of distortion of the recognition site. The disulfide bridges could be cleaved using a large excess of DTT to regenerate the corresponding sulfhydryl compounds. A study of the serum stabilities of disulfide-bridged oligonucleotides showed that the bridged duplexes were much more stable than their unmodified counterparts, whereas the rate of degradation of the stem-loop structures was more dependent upon the size of the loop than the presence or absence of the disulfide bridge. In summary, we have described a novel methodology, employing commercially available reagents, for the stabilization of oligonucleotide duplexes or stem-loop structures by disulfide bridge formation. 相似文献
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S. K. Katti A. H. Robbins Y. Yang W. W. Wells 《Protein science : a publication of the Protein Society》1995,4(10):1998-2005
We report here the first three-dimensional structure of a mammalian thioltransferase as determined by single crystal X-ray crystallography at 2.2 A resolution. The protein is known for its thiol-redox properties and dehydroascorbate reductase activity. Recombinant pig liver thioltransferase expressed in Escherichia coli was crystallized in its oxidized form by vapor diffusion technique. The structure was determined by multiple isomorphous replacement method using four heavy-atom derivatives. The protein folds into an alpha/beta structure with a four-stranded mixed beta-sheet in the core, flanked on either side by helices. The fold is similar to that found in other thiol-redox proteins, viz. E. coli thioredoxin and bacteriophage T4 glutaredoxin, and thus seems to be conserved in these functionally related proteins. The active site disulfide (Cys 22-Cys 25) is located on a protrusion on the molecular surface. Cys 22, which is known to have an abnormally low pKa of 3.8, is accessible from the exterior of the molecule. Pro 70, which is in close proximity to the disulfide bridge, assumes a conserved cis-peptide configuration. Mutational data available on the protein are in agreement with the three-dimensional structure. 相似文献
136.
催产素在脊髓水平对电针镇痛的影响 总被引:3,自引:0,他引:3
采用玻璃微电极胞外记录和脊髓表面给药的方法观察了催产素(OT)、抗催产素血清(AOTS)以及电针穴位对背角神经元伤害性诱发放电的影响。结果表明:电针穴位或脊髓表面施加OT可部分抑制脊髓背角神经元的伤害性诱发放电;在电针的基础上施加OT则明显加强电针的抑制效应;相反,用AOTS预处理后,电针的抑制作用放取消。提示OT在脊髓水平参与了对痛觉信息的调制,并与一定频率的针刺镇痛有关。 相似文献
137.
Specific absorption rates (SARs) were determined theoretically and experimentally for several spherical models of tissue exposed to electrical fields of TE101 mode in a rectangular cavity of 57.3 MHz resonant frequency. The approximate theoretical SAR can be calculated according to the Mie theory by superposition of four plane waves representing the fields excited in the cavity. The theoretical and thermographically determined SAR patterns in spheres with radii of 5, 7.5, and 10 cm and with conductivities of 0.1, 1, and 10 S/m were compared. For a sphere with radius less than 7.5 cm and conductivity less than 1 S/m, the SAR was quite uniform. When conductivity was increased to 10 S/m, the SAR patterns showed higher absorption in the periphery of the largest sphere (10-cm radius). These characteristics are important in evaluating the scaling technique of exposing a model of a human to very-high-frequency fields to obtain power absorption data in humans exposed to high-frequency or very-low-frequency fields. 相似文献
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