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31.
32.
Possible Involvement of Phosphorylation of Occludin in Tight Junction Formation 总被引:21,自引:3,他引:18 下载免费PDF全文
Akira Sakakibara Mikio Furuse Mitinori Saitou Yuhko Ando-Akatsuka Shoichiro Tsukita 《The Journal of cell biology》1997,137(6):1393-1401
Occludin is an integral membrane protein localizing at tight junctions in epithelial and endothelial cells. Occludin from confluent culture MDCK I cells resolved as several (>10) bands between 62 and 82 kD in SDS-PAGE, of which two or three bands of the lowest Mr were predominant. Among these bands, the lower predominant bands were essentially extracted with 1% NP-40, whereas the other higher Mr bands were selectively recovered in the NP-40–insoluble fraction. Alkaline phosphatase treatment converged these bands of occludin both in NP-40–soluble and -insoluble fractions into the lowest Mr band, and phosphoamino acid analyses identified phosphoserine (and phosphothreonine weakly) in the higher Mr bands of occludin. These findings indicated that phosphorylation causes an upward shift of occludin bands and that highly phosphorylated occludin resists NP-40 extraction. When cells were grown in low Ca medium, almost all occludin was NP-40 soluble. Switching from low to normal Ca medium increased the amount of NP-40–insoluble occludin within 10 min, followed by gradual upward shift of bands. This insolubilization and the band shift correlated temporally with tight junction formation detected by immunofluorescence microscopy. Furthermore, we found that the anti–chicken occludin mAb, Oc-3, did not recognize the predominant lower Mr bands of occludin (non- or less phosphorylated form) but was specific to the higher Mr bands (phosphorylated form) on immunoblotting. Immunofluorescence microscopy revealed that this mAb mainly stained the tight junction proper of intestinal epithelial cells, whereas other anti-occludin mAbs, which can recognize the predominant lower Mr bands, labeled their basolateral membranes (and the cytoplasm) as well as tight junctions. Therefore, we conclude that non- or less phosphorylated occludin is distributed on the basolateral membranes and that highly phosphorylated occludin is selectively concentrated at tight juctions as the NP-40–insoluble form. These findings suggest that the phosphorylation of occludin is a key step in tight junction assembly. 相似文献
33.
Movements of organelles in the nuclear region as the cell cycleprogresses in single-celled protonemata of Adiantum capillus-veneriswere examined by digital image processing techniques and microscopyof particle movement. Organelles in the nuclear region werenot very crowded and moving directionally along the longitudinalaxis of the filamentous cell in the G1 and S phases. They beganto gather and accumulate in the nuclear region in early G2 phase,after which directional movement changed to undirectional Brownianmotion-like movement in late G2 phase. Movement of organelleslocated on the lateral surface of the nucleus slowed after premitoticpositioning of nucleus and lasted until the nucleolus disappeared.Movement of organelles in the cytoplasm surrounding the nucleoplasmresumed just after the nucleolus disappeared, whereas organelleslocated in the outer regions of the apical and basal surfacesof the nucleus moved rapidly during prophase but did not moveduring metaphase, movement being resumed after chromosome separation.Thus, organelle movement in the nuclear region showed temporaland spatial change during the cell cycle. (Received August 24, 1983; Accepted December 28, 1983) 相似文献
34.
Ichiro Azuma Mikio Yamawaki Kosei Yasumoto Yuichi Yamamura 《Cancer immunology, immunotherapy : CII》1978,4(2):95-100
Summary The antitumor activity of the cell wall skeleton preparations of four species of Nocardia, N. brasiliensis strain 146, N. coeliaca strain 122, N. polychromogenes strain 6, and N. rubra, which showed potent adjuvant activity on the induction of cell-mediated cytotoxicity in allogeneic mice, was examined with the aid of EL-4 leukemia, melanoma B16, and MH-134 hepatoma in syngeneic mice. Preliminary clinical trials were performed and the results suggest that the cell wall skeleton of N. rubra, upon intrapleural injection, may be useful as an immunotherapeutic agent for patients with malignant pleurisy. The chemical properties of these cell wall skeleton preparations are described. 相似文献
35.
Mikio Kobayashi 《Journal of plant research》1977,90(2):143-148
The proliferation patterns of the green alga,Ankistrodesmus gracilis were investigated by electron microscopy. The multichotomically dividing phase, namely the type I phase was characterized
by multichotomical nuclear division, whereas the dichotomically dividing phase, i.e., the type II phase was related to dichotomical
nuclear division. Type I phase was controlled by a lower cell density and a diurnal light-dark rhythm of LD 14:10. Type II
phase was induced by a cell density of more than 1.0×107 cells/ml. Independent of the 2 types of proliferation patterns, the diurnal light-dark rhythms related to the autospore release from
a mother cell. The diurnal light-dark rhythm of LD 14∶10 induced the scattered release of autospores. On the other hand, a
rhythm of LD 18∶6 induced the release of a couple of tightly adhered autospores from a mother cell which had been observed
apparently as a type II cell by light microscopy. 相似文献
36.
The effects of various kinds of nitrogen compounds on the proliferation patterns ofAnkistrodesmus gracilis were examined. Among the tested compounds, the addition of peptone, arginine or glutamine to the N-free medium was the most effective on cell growth. The effectiveness was also shown in media containing one of several kinds of amino acid. In both peptone and arginine media the alga proliferated with multiple fission type of cell division, whereas the cultures grown in glutamine or serine medium contained 2-celled colonies at high frequencies. The latter was an efficient culture condition for cell reproduction with binary fission. The evidence for the behavior of nuclear and cytoplasmic division in these growth patterns were obtained from observations of thin sectioned materials. 相似文献
37.
Summary A serum-free culture system supplemented with neural tissue extract for normal and tumor human esophagi was applied to the
culture of mouse esophageal epithelium. Similar to mouse mesenchyme and skin epithelium, esophageal epithelial lines (MEE)
emerged after serial culture. The cells had an apparent unlimited life span but retained morphology and other characteristics
of normal epithelial cells. The cells formed a small cyst consisting of keratined squamous epithelium in syngenic hosts. A
screen for growth factors that stimulated growth of the nonmalignant MEE cells in the absence of neural extract revealed that
epidermal growth factor (EGF) and heparin-binding (fibroblast) growth factors (HBGF) were most effective. An HBGF-like activity
was apparent in extracts of rapidly proliferating but not quiescent MEE cells at low or confluent densities. A cloned cell
line (MEE/C8) was selected from MEE cell cultures in the absence of neural extract. MEE/C8 cells proliferated independent
of either EGF or HBGF at rates equal to MEE cells, cell extracts exhibited HBGF-like activity at all stages of proliferation,
and the cells formed large invasive tumors in syngenic hosts. The HBGF-like activity present in extracts of tumorigenic MEE/C8
and proliferating nonmalignant MEE cells had properties similar to HBGF-1 (acidic fibroblast growth factor). These results
constitute a cultured mouse esophageal epithelial cell model for study of conversion of immortalized premalignant cells to
malignant cells, and suggest that conversion from a state of cell cycle-dependent autocrine expression of one or more members
of the HBGF family to a state of constitutive expression correlates with and may contribute to malignancy.
The work was supported in part by grants CA37589 and DK35310 to Dr. McKeehan, from the National Cancer Institute, Bethesda,
MD. 相似文献
38.
Shinji Kakudo Kazumasa Yoshikawa Mikio Tamaki Etsuo Nakamura Hiroshi Teraoka 《Applied microbiology and biotechnology》1992,38(2):226-233
Summary In order to obtain a large quantity of glutamic-acid-specific endopeptidase of Staphylococcus aureus ATCC12600 (SPase) without cultivating its pathogenic host bacterium, expression plasmids enabling secretion of SPase from Bacillus subtilis were constructed by inserting the SPase gene into B. subtilis-Escherichia coli shuttle vectors. B. subtilis harbouring a simple recombinant plasmid containing the coding and the 5-flanking regions of SPase in the shuttle vector pHY300PLK secreted 22 mg/l of SPase into the medium. As this level was lower than that of the natural strain (45 mg/l), we tried to increase the expression level by constructing a series of hybrid plasmids with the following features: (1) the terminator sequence of the alkaline protease gene from B. subtilis, (2) the promoter and the leader sequences of the -amylase gene or of alkaline protease gene from B. amyloliquefaciens, (3) the vector pHY300PLK and the fused vector of pHY300PLK and pUB110. By using a variety of hybrid plasmids, the resulting transformants secreted SPase at levels of 33–120 mg/l. The recombinant SPase isolated from the medium was indistinguishable from the natural one with respect to its behaviour on sodium dodecyl sulphate-polyacrylamide gel electrophoresis and Western blotting as well as its enzyme activity.Correspondence to: S. Kakudo 相似文献
39.
Jun Ishizaki Mikio Tamaki Masaru Shin Hiroshige Tsuzuki Kazumasa Yoshikawa Hiroshi Teraoka Nobuo Yoshida 《Applied microbiology and biotechnology》1992,36(4):483-486
Summary Recombinant human glucagon was succesfully produced with a high level of expression in Escherichia coli as a fusion protein with human interferon . The synthetic gene was designed to release glucagon, which does not contain glutamic acid residues, from fusion protein with the Staphylococcus aureus strain V8 protease that specifically cleaves the peptide bond on the carboxyl side of the glutamic acid residue. The resulting glucagon was purified to homogeneity by a combination of C18 reverse-phase HPLC and ion-exchange HPLC. The yield of intact glucagon obtained from 11 of culture was approximately 12 mg. The structure of recombinant human glucagon was confirmed by HPLC and amino acid composition/sequence analyses.
Offprint requests to: J. Ishizaki 相似文献
40.
Distribution of ribonucleic acid coliphages in raw sewage from treatment plants in Japan. 总被引:4,自引:2,他引:2 下载免费PDF全文
To determine the transmission cycle of ribonucleic acid (RNA) coliphages in their natural habitats, we investigated the distribution patterns of RNA phages in raw sewage collected from treatment plants in various localities in Japan. Most of the sewage samples contained group II and III phages. Samples from treatment plants in Sapporo, Tokyo, and Toyama contained appreciable amounts of group I phages in addition to the group II and III phages. As a whole, raw sewage from treatment plants in Japan contained RNA phages of the three groups in the ratio 1:2:5, group I/II/III. Based on the distribution patterns of RNA phages in sewage from domestic drainage in Japan proper (group II/III, 3:1), in animal feces and sewage from slaughter houses (mostly group I), and in human feces (group II/III, 1:1), it can be reasonably said that group I phages tend to be introduced from animal sources and group II and III phages tend to be introduced from human sources. Raw sewage from treatment plants in Japan consists mainly of human feces, sewage from domestic drainage, and industrial wastewater, and, in part, from slaughter houses. In fact, sewage from slaughter houses together with that from human sources flowed into the treatment plants of Tokyo as far as we could confirm. 相似文献