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991.
Two DNA-dependent DNA-polymerases (E.C. 2.7.7.7) are partially purified from the high speed supernatant of mechanically disrupted hyphae of Neurospora crassa WT 74A. Some properties such as temperature and pH optimum and theoptimal concentrations for Mg2+, Zn2+, NH4+ and Na+ are very similar. On the other hand these enzymes show different properties on ion-exchange columns, are well distinguished by molecular weight (147 000 d and 110 000 d for A and B resp.) and the stimulation by K+ differs (K+ optimum for A: 5-70 mM and for B: 45 mM). Mn2+ and Zn2+ inhibit incorporation of deoxyribonucleoside monophosphates between 70 and 90%. Our best preparations so far have specific activities of 13 200 units/mg protein for A and 12 000 units/mg protein for B. 相似文献
992.
The molecular chaperone SecB is released from the carboxy-terminus of SecA during initiation of precursor protein translocation. 总被引:11,自引:1,他引:11 下载免费PDF全文
The chaperone SecB keeps precursor proteins in a translocation-competent state and targets them to SecA at the translocation sites in the cytoplasmic membrane of Escherichia coli. SecA is thought to recognize SecB via its carboxy-terminus. To determine the minimal requirement for a SecB-binding site, fusion proteins were created between glutathione-S-transferase and different parts of the carboxy-terminus of SecA and analysed for SecB binding. A strikingly short amino acid sequence corresponding to only the most distal 22 aminoacyl residues of SecA suffices for the authentic binding of SecB or the SecB-precursor protein complex. SecAN880, a deletion mutant that lacks this highly conserved domain, still supports precursor protein translocation but is unable to bind SecB. Heterodimers of wild-type SecA and SecAN880 are defective in SecB binding, demonstrating that both carboxy-termini of the SecA dimer are needed to form a genuine SecB-binding site. SecB is released from the translocase at a very early stage in protein translocation when the membrane-bound SecA binds ATP to initiate translocation. It is concluded that the SecB-binding site on SecA is confined to the extreme carboxy-terminus of the SecA dimer, and that SecB is released from this site at the onset of translocation. 相似文献
993.
C Masutani M Araki K Sugasawa P J van der Spek A Yamada A Uchida T Maekawa D Bootsma J H Hoeijmakers F Hanaoka 《Molecular and cellular biology》1997,17(12):6915-6923
hHR23B was originally isolated as a component of a protein complex that specifically complements nucleotide excision repair (NER) defects of xeroderma pigmentosum group C cell extracts in vitro and was identified as one of two human homologs of the Saccharomyces cerevisiae NER gene product Rad23. Recombinant hHR23B has previously been shown to significantly stimulate the NER activity of recombinant human XPC protein (rhXPC). In this study we identify and functionally characterize the XPC-binding domain of hHR23B protein. We prepared various internal as well as terminal deletion products of hHR23B protein in a His-tagged form and examined their binding with rhXPC by using nickel-chelating Sepharose. We demonstrate that a domain covering 56 amino acids of hHR23B is required for binding to rhXPC as well as for stimulation of in vitro NER reactions. Interestingly, a small polypeptide corresponding to the XPC-binding domain is sufficient to exert stimulation of XPC NER activity. Comparison with known crystal structures and analysis with secondary structure programs provided strong indications that the binding domain has a predominantly amphipathic alpha-helical character, consistent with evidence that the affinity with XPC is based on hydrophobic interactions. Our work shows that binding to XPC alone is required and sufficient for the role of hHR23B in in vitro NER but does not rule out the possibility that the protein has additional functions in vivo. 相似文献
994.
Jouke P. Kardolus Herman J. van Eck Ronald G. van den Berg 《Plant Systematics and Evolution》1998,210(1-2):87-103
Using the AFLP technique highly informative DNA fingerprints were generated from 19 taxa ofSolanum sect.Petota (potatoes) and three taxa ofSolanum sect.Lycopersicum (tomatoes). Both phenetic and cladistic analyses were conducted from the individual genotypic level to the species level. An AFLP fingerprint, using a combination of suitable AFLP primers, generated 12 to 71 scorable fragments per genotype which was sufficient for taxonomic interpretation. The classifications based on the molecular markers were generally in agreement with current taxonomic opinions. Unexpectedly,S. microdontum was associated with ser.Megistacroloba rather than with ser.Tuberosa, andS. demissum (ser.Demissa) and species of ser.Acaulia appeared closely affiliated. AFLP is an efficient and reliable technique to generate biosystematic data and therefore a promising tool for evolutionary studies. 相似文献
995.
G.J.H. Buisman C.T.W. van Helteren G.F.H. Kramer J.W. Veldsink J.T.P Derksen F.P. Cuperus 《Biotechnology letters》1998,20(2):131-136
Of seven lipases used to esterify functionalized phenols and fatty acids to synthesize lipophilic antioxidants, that from Candida antarctica lipase B (CAL-B) had the highest catalytic activity. Esterification yields of benzoic acid derivatives catalyzed by CAL-B are below 2% after 7 days. In contrast, relatively high yields were obstained for the esterification of (poly)phenols bearing primary hydroxy groups and aliphatic acids (85% yield within 15 hours). Crude products were purified and used as lipophilic antioxidants in sunflower oil. 相似文献
996.
J. A. Wilmer S. R. Abrams J. P. F. G. Helsper L. H. W. van der Plas 《Journal of Plant Growth Regulation》1998,17(1):19-23
Modification of the structure of abscisic acid (ABA) has been reported to result in modification of its physiologic activity.
In this study we tested the effect of removing methyl groups from the ring and of chirality of ABA on activity in microspore-derived
embryos of oilseed rape (Brassica napus L.). The natural (+)-ABA molecule induced growth inhibition and an increase in the amount of erucic acid accumulated in the
oil at medium concentrations less than 1 μm. (−)-ABA showed similar effects. Removing the 7′-methyl group resulted in a dramatic decrease in activity: (+)-7′-demethyl-ABA
retained some activity as a growth inhibitor; a 10–100 μm concentration of this compound was needed for a response, and (−)-7′-demethyl-ABA was almost completely inactive. Similar
effects were observed with regard to elongase activity, which catalyzes erucic acid biosynthesis from oleic acid. Removal
of the 8′- and 9′-methyl groups resulted in a more complex response. These compounds all showed intermediate activity; for
growth inhibition, the presence of the 9′-methyl was the more important determinant, whereas chirality dominated the response
on erucic acid accumulation, with the (+)-enantiomers being more active.
Received July 25, 1997; accepted October 31, 1997 相似文献
997.
Krook J; Vreugdenhil D; Dijkema C; van der Plas L 《Journal of experimental botany》1998,49(329):1917-1924
Cells were grown in batch culture on a mixture of 50 mM glucose and
fructose as the carbon source; either the glucose or the fructose was
[1-13C]-labelled. In order to investigate the uptake
and conversion of glucose and fructose during long-term labelling
experiments in cell suspensions of Daucus carota L.,
samples were taken every 2 d during a 2 week culture period and sucrose and
starch were assayed by means of HPLC and 13C-nuclear
magnetic resonance. The fructose moieties of sucrose had a lower labelling
percentage than the glucose moieties. Oxidative pentose phosphate pathway
activity in the cytosol is suggested to be responsible for this loss of
label of especially C-1 carbons. A combination of oxidative pentose
phosphate pathway activity, a relatively high activity of pathway to
sucrose synthesis and a slow equilibration between glucose-6-phosphate and
fructose-6-phosphate could explain these results. Starch contained glucose
units with a much lower labelling percentage than glucose moieties of
sucrose: it was concluded that a second, plastid-localized, oxidative
pentose phosphate pathway was responsible for removal of C-1 carbons of the
glucosyl units used for synthesis of starch. Redistribution of label from
[1-13C]-hexoses to
[6-13C]-hexoses also occurred: 18-45% of the label
was found at the C-6 carbons. This is a consequence of cycling between
hexose phosphates and those phosphates in the cytosol catalysed by PFP. The
results indicate that independent (oxidative pentose phosphate pathway
mediated) sugar converting cycles exist in the cytosol and
plastid.Key words: Daucus carotaL., cell suspensions,
carbon-13 nuclear magnetic resonance, 13C-NMR,
carbohydrate cycling, oxidative pentose phosphate pathway, plastid.
相似文献
998.
L. Barton Browne A. C. M. van Gerwen W. G. Vogt 《Entomologia Experimentalis et Applicata》1990,55(1):33-40
Oviposition by Lucilia cuprina Wiedemann (Diptera, Calliphoridae) was examined in relation to period of oviposition site-deprivation and egg-load. Effects of oviposition site-deprivation were examined by comparing oviposition performance of individual females that had matured their batch of oocytes within the previous 24 h with that of females which had reached ovarian maturity 8 days previously. Egg-load was manipulated by causing females of this anautogenous species to consume different amounts of protein-rich material. In no-choice experiments, individual females of the different categories were given access for 4 h to oviposition substrate, soaked with (i) liver exudate, (ii) the exudate diluted 16-fold or (iii) the undilated exudate containing the oviposition deterrent sodium chloride at a concentration of 2 M. These solutions elicited oviposition from different proportions of females, but neither these proportions, nor the interval between introduction of the oviposition site and the initiation of oviposition, was significantly affected by the period of oviposition site-deprivation or the number of eggs matured by the females.
Résumé L'effet de la privation de lieu de ponte a été étudié en comparant les pontes de femelles isolées ayant formé leurs ufs mûrs dans les 24 heures précédentes, à celles de femelles ayant atteint leur maturité sexuelle 8 jours avant. La rétention ovocytaire est provoquée en faisant consommer aux femelles de cette espèce anautogène différentes quantités d'aliments riches en protéines. La ponte de femelles dont le contingent total de leurs ovocytes s'est développé, — c'est-à-dire 260 —, après consommation ad libitum de foie de mouton pendant 48 heures, a été comparée à celle de femelles ayant formé 190 ovocytes mûrs après ingestion d'une quantité limitée de jus de foie.Dans des expériences sans choix, les femelles isolées de différences catégories ont eu accès pendant 4 heures au substrat de ponte trempé: 1) dans du jus de foie, 2) dans du jus dilué 16 fois, 3) dans du jus de foie non dilué mais contenant NaCl (inhibiteur de la ponte) à la concentration de 2 M. Le jus non dilué a provoqué une forte stimulation, induisant la ponte de 80% des femelles. Le jus dilué et celui contenant NaCl n'ont induit la ponte que de 40% des femelles avec des niveaux de stimulation bien plus faibles. La date d'introduction du lieu de ponte et le taux de rétention des ovocytes mûrs n'ont eu auçun effet sur la proportion de femelles réagissant à ces 3 types de stimulation.相似文献
999.
We have recently described an insulin-resistant patient with leprechaunism (leprechaun G.) having a homozygous leucine----proline mutation at amino acid position 233 in the alpha-chain of the insulin receptor. The mutation results in a loss of insulin binding to cultured fibroblasts. Fibroblasts from the patient and control individuals were used to quantify the stimulation of 2-deoxyglucose uptake by insulin and insulin-like growth factor 1 (IGF-1). Insulin hardly stimulates basal 2-deoxyglucose uptake in the patient's fibroblasts whereas in control fibroblasts the uptake of 2-deoxyglucose is stimulated by insulin approximately 1.7 times. In contrast, IGF-1 stimulates hexose uptake in the patient's fibroblasts 1.8 times, a similar value to that obtained by stimulation of control fibroblasts with insulin or IGF-1. With both types of fibroblasts, maximal IGF-1 response is reached at about 10 nM IGF-1, the ED50 being approximately 4 nM. The results indicate that the insulin responsive glucose transport in primary fibroblasts is functionally linked to the receptor for IGF-1. Insulin binds with an approximately 200-fold lower affinity to IGF-1 receptors, compared to homologous IGF-1 binding. As an insulin concentration of 10 microM is unable to give maximal stimulation of glucose uptake in the patient's fibroblasts, which is already seen with 10 nM IGF-1, it seems that occupation of IGF-1 receptors by insulin on the patient's cells is less efficient at stimulating hexose uptake compared to homologous activation. 相似文献
1000.
M. Koornneef T. D. G. Bosma C. J. Hanhart J. H. van der Veen J. A. D. Zeevaart 《TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik》1990,80(6):852-857
Summary In tomato, nine independent EMS-induced mutants representing recessive mutations at three different loci (gib-1, gib-2, and gib-3) were isolated. Six of these have an almost absolute gibberellin requirement for seed germination and elongation growth. In addition, the leaves are darker green, smaller, and changed in structure as compared to wild type. The three other mutants, which germinate without GA, are allelic to specific, nongerminating mutants and have less severe mutant characteristics. The respective loci are situated on three different chromosomes. The genes identified by these mutants control steps in gibberellin biosynthesis, as endogenous gibberellins are strongly reduced. 相似文献