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Calcium regulates inositol 1,3,4,5-tetrakisphosphate production in lysed thymocytes and in intact cells stimulated with concanavalin A. 总被引:2,自引:0,他引:2 下载免费PDF全文
Lysed mouse thymocytes release [3H]inositol 1,4,5 trisphosphate from [3H]inositol-labelled phosphatidyl inositol 4,5-bisphosphate in response to GTP gamma S, and rapidly phosphorylate [3H]inositol 1,4,5-trisphosphate to [3H]inositol 1,3,4,5-tetrakisphosphate. The rate of phosphorylation is increased approximately 7-fold when the free [Ca2+] in the lysate is increased from 0.1 to 1 microM, the range in which the cytosolic free [Ca2+] increases in intact thymocytes in response to the mitogen concanavalin A. Stimulation of the intact cells with concanavalin A also results in a rapid and sustained increase in the amount of inositol 1,3,4,5-tetrakisphosphate, and a much smaller transient increase in 1,4,5-trisphosphate. Lowering [Ca2+] in the medium from 0.4 mM to 0.1 microM before addition of concanavalin A reduces accumulation of inositol 1,3,4,5-tetrakisphosphate by at least 3-fold whereas the increase in inositol 1,4,5-trisphosphate is sustained rather than transient. The data imply that in normal medium the activity of the inositol 1,4,5-trisphosphate kinase increases substantially in response to the rise in cytosolic free [Ca2+] generated by concanavalin A, accounting for both the transient accumulation of inositol 1,4,5-trisphosphate and the sustained high levels of inositol 1,3,4,5-tetrakisphosphate. Inositol 1,3,4,5-tetrakisphosphate is a strong candidate for the second messenger for Ca2+ entry across the plasma membrane. This would imply that the inositol polyphosphates regulate both Ca2+ entry and intracellular Ca2+ release, with feedback control of the inositol polyphosphate levels by Ca2+. 相似文献
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T. P. Wallace A. C. Stewart D. Pappin C. J. Howe 《Molecular & general genetics : MGG》1989,216(2-3):334-339
Summary A 9 kDa polypeptide which is loosely attached to the inner surface of the thylakoid membrane and is important for the oxygen-evolving activity of Photosystem II in the thermophilic cyanobacterium Phormidium laminosum has been purified, a partial amino acid sequence obtained and its gene cloned and sequenced. The derived amino acid sequence indicates that the 9 kDa polypeptide is initially synthesised with an N-terminal leader sequence of 44 amino acids to direct it across the thylakoid membrane. The leader sequence consists of a positively charged N-terminal region, a long hydrophobic region and a typical cleavage site. These features have analogous counterparts in the thylakoid-transfer domain of lumenal polypeptides from chloroplasts of higher plants. These findings support the view of the proposed function of this domain in the two-stage processing model for import of lumenal, nuclear-encoded polypeptides. In addition, there is striking primary sequence homology between the leader sequences of the 9 kDa polypeptide and those of alkaline phosphatase (from the periplasmic space of Escherichia coli) and, particularly in the region of the cleavage site, the 16 kDa polypeptide of the oxygen-evolving apparatus in the thylakoid lumen of spinach chloroplasts. 相似文献
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PD Dr. G. F. Jirikowski J. F. Ramalho-Ortigao K. W. Kesse F. E. Bloom 《Histochemistry and cell biology》1990,94(2):187-190
Summary We recently described a nonradioactive method for in situ hybridization with 5-bromo-2-deoxyuridine (BrdU) labelled oligonucleotide
probes. An antibody to BrdU and immunocytochemistry were used in order to detect the hybridization signal. We have now applied
this method to semithin Epon sections, in order to hybridize consecutive sections through single cells with different probes
and to stain them with antibodies to neuropeptides. It could be shown that Epon embedding preserves mRNA well. In the present
study we used a BrdU labelled synthetic oligonucleotide probe complementary to a fragment of the vasopressin precursor and
an antibody to Arg-vasopressin. Vasopressin mRNA was demonstrable in a fraction of the vasopressin immunoreactive neurons
in the magnocellular nuclei. In addition some of the magnocellular neurons showed either hybridization or vasopressin immunostaining
only, perhaps indicating different stages of synthetic and secretory activity.
The method described seems to be a valuable tool for studying synthetic activity in peptidergic neurons on a single cell level.
The method might also have potential for in situ hybridization on the electronmicroscopical level. 相似文献
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Transforming growth factor beta 1 inhibition of p34cdc2 phosphorylation and histone H1 kinase activity is associated with G1/S-phase growth arrest. 总被引:29,自引:8,他引:21 下载免费PDF全文
Transforming growth factor beta 1 (TGF beta 1) is a potent inhibitor of epithelial cell proliferation. We present data which indicate that epithelial cell proliferation is inhibited when TGF beta 1 is added throughout the prereplicative G1 phase. Cultures become reversibly blocked in late G1 at the G1/S-phase boundary. The inhibitory effects of TGF beta 1 on cell growth occur in the presence of the RNA synthesis inhibitor 5,6-dichloro-1-beta-D-ribofuranosylbenzimidazole. Associated with this inhibitory effect is a decrease in the phosphorylation and histone H1 kinase activity of the p34cdc2 protein kinase. These data suggest that TGF beta 1 growth inhibition in epithelial cells involves the regulation of p34cdc2 activity at the G1/S transition. 相似文献
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Functional status of interleukin 2 receptors expressed by immature (Lyt-2-/L3T4-) thymocytes 总被引:9,自引:0,他引:9
J W Lowenthal R C Howe R Ceredig H R MacDonald 《Journal of immunology (Baltimore, Md. : 1950)》1986,137(8):2579-2584
A subpopulation of phenotypically immature (Lyt-2-/L3T4-) thymocytes express receptors for the polypeptide hormone interleukin 2 (IL 2); however, these cells do not proliferate in vitro in response to IL 2. In investigating this phenomenon in greater detail, we observed that the IL 2 receptors (IL 2-R) on freshly isolated immature thymocytes bound IL 2 with about fivefold lower affinity (Kd approximately 100 pM) than IL 2-R on activated mature T cells and T cell lines (Kd approximately 20 pM). Furthermore, in contrast to activated T cells, Lyt-2-/L3T4- thymocytes did not endocytose bound IL 2. When stimulated in short-term culture with a combination of phorbol ester (PMA) and calcium ionophore, Lyt-2-/L3T4- thymocytes proliferated in a largely IL 2-dependent fashion. IL 2-R expression on these activated cells initially disappeared (at 24 hr) and subsequently reappeared (at 48 to 72 hr). Reexpressed IL 2-R on activated thymocytes resembled those on mature T lymphocytes in that they bound IL 2 with high affinity (Kd = 15 to 25 pM) and were capable of endocytosing IL 2. Taken together, these data place certain constraints on the putative physiologic role of IL 2 in intrathymic growth regulation. 相似文献