全文获取类型
收费全文 | 4405篇 |
免费 | 318篇 |
国内免费 | 2篇 |
出版年
2023年 | 18篇 |
2022年 | 38篇 |
2021年 | 78篇 |
2020年 | 36篇 |
2019年 | 52篇 |
2018年 | 104篇 |
2017年 | 101篇 |
2016年 | 154篇 |
2015年 | 210篇 |
2014年 | 217篇 |
2013年 | 275篇 |
2012年 | 322篇 |
2011年 | 322篇 |
2010年 | 224篇 |
2009年 | 193篇 |
2008年 | 226篇 |
2007年 | 249篇 |
2006年 | 207篇 |
2005年 | 198篇 |
2004年 | 179篇 |
2003年 | 135篇 |
2002年 | 139篇 |
2001年 | 109篇 |
2000年 | 63篇 |
1999年 | 58篇 |
1998年 | 45篇 |
1997年 | 24篇 |
1996年 | 30篇 |
1995年 | 32篇 |
1994年 | 26篇 |
1993年 | 25篇 |
1992年 | 40篇 |
1991年 | 25篇 |
1990年 | 37篇 |
1989年 | 36篇 |
1988年 | 27篇 |
1987年 | 36篇 |
1986年 | 19篇 |
1985年 | 22篇 |
1984年 | 24篇 |
1983年 | 21篇 |
1982年 | 19篇 |
1981年 | 18篇 |
1977年 | 21篇 |
1976年 | 22篇 |
1975年 | 27篇 |
1974年 | 19篇 |
1973年 | 20篇 |
1971年 | 17篇 |
1966年 | 17篇 |
排序方式: 共有4725条查询结果,搜索用时 15 毫秒
51.
Scale-up of the adenovirus expression system for the production of recombinant protein in human 293S cells 总被引:6,自引:0,他引:6
Alain Garnier Johanne Côté Isabelle Nadeau Amine Kamen Bernard Massie 《Cytotechnology》1994,15(1-3):145-155
Human 293S cells, a cell line adapted to suspension culture, were grown to 5×106 cells/mL in batch with calcium-free DMEM. These cells, infected with new constructions of adenovirus vectors, yielded as much as 10 to 20% recombinant protein with respect to the total cellular protein content. Until recently, high specific productivity of recombinant protein was limited to low cell density infected cultures of no more than 5×105 cells/mL. In this paper, we show with a model protein, Protein Tyrosine Phosphatase 1C how high product yield can be maintained at high cell densities of 2×106 cells/mL by a medium replacement strategy. This allows the production of as much as 90 mg/L of active recombinant protein per culture volume. Analysis of key limiting/inhibiting medium components showed that glucose addition along with pH control can yield the same productivity as a medium replacement strategy at high cell density in calcium-free DMEM. Finally, the above results were reproduced in 3L bioreactor suspension culture thereby establishing the scalability of this expression system. The process we developed is used routinely with the same success for the production of various recombinant proteins and viruses.Abbreviations CFDMEM
calcium-free DMEM
- CS
bovine calf serum
- hpi
hours post-infection
- J+
enriched Joklik medium
- MLP
major late promoter
- MOI
multiplicity of infection (# of infectious viral particle/cell)
- q
specific consumption rate (mole/cell.h)
- pfu
plaque forming unit (# of infectious viral particle)
- Y
yield (g/E6 cells or mole/cell) 相似文献
52.
Marie J. Richard Veronique Ducros Michel Rorêt Josiane Arnaud Charles Coudray Michèle Fusselier Alain Favier 《Biological trace element research》1993,39(2-3):149-159
In six chronic dialyzed uremic patients, an intravenous sodium selenite (Se 50 μg during 5 wk and then 100 μg) and zinc gluconate
(Zn 5 mg) supplementation was performed during 20 wk at each dialysis session three times weekly. Before supplementation,
plasma Se and Zn, plasma and erythrocytes (RBC) antioxidant metalloenzymes glutathione peroxidase (GPX), and superoxide dismutase
(SOD) were significantly decreased, whereas lipid peroxidation (as thiobarbituric acid reactants TBARs) was increased. To
obtain a significative change in plasma selenium, we had to use an Se dose of 100 μg/dialysis session. Then, treatment-increased
plasma Se (from 0.58 ±0.09 to 0.89±0.16 μmol/L) led to a repletion of RBC-GPX (from 29.6±6 to 43±5.8 U/g Hb) and increased
plasma GPX levels (from 62±13 to 151±43 U/L). Plasma Zn and RBC-SOD did not vary significantly. The change of TBARs was not
observed between wk 1 and 4. They decreased significantly between wk 4 (4.80±0.21μmol/L) and wk 20 (4.16±0.26 μmol/L). We
noted a low correlation between TBARs and plasma GPX. A strong correlation was observed between Se and plasma GPX. The reversal
of Se deficiencies should reduce oxidative damage observed in these patients. 相似文献
53.
Biochemical profiles on API Rapid CH* strips and protein profiles on polyacrylamide gels in the presence of sodium dodecyl sulfate were used to distinguish two
strains of the entomopathogenic fungusBeauveria bassiana (Balsamo) Vuillemin, ARSEF 2991 and ATCC 44860. Next, the toxicity of these two strains was determined at concentrations
of 102, 104, 106 and 108 blastospores/ml on larvae of the Colorado potato beetleLeptinotarsa decemlineata Say (Coleoptera: Chrysomelidae) and of its predator, the spotted ladybird beetle,Coleomegilla maculata lengi Timberlake (Coleoptera: Coccinellidae).
Both strains were highly toxic toL. decemlineata larvae. However, the two strains exhibited different levels of toxicity forC. maculata larvae: ARSEF 2991 was toxic, whereas ATCC 44860 caused little coccinellid larval mortality.
Résumé Les profils biochimiques sur galeries API Rapid CH* et les profils protéiques sur gels de polyacrylamide ont été utilisés pour distinguer deux souches du champignon entomopathogèneBeauveria bassiana (Balsamo) Vuillemin. La toxicité de ces deux souches a été déterminée à des concentrations de 102, 104, 106 et 108 blastospores/ml sur des larves du doryphore,Leptinotarsa decemlineata Say (Coleoptera: Chrysomelidae) et de la coccinelle maculéeColeomegilla maculata lengi Timberlake (Coleoptera: Coccinellidae). Les deux souches deB. bassiana se sont avérées actives à l'égard des larves deL. decemlineata. Toutefois la souche ARSEF 2991 s'est avérée pathogène pour les larves deC. maculata, alors que la souche ATCC 44860 a provoqué une faible mortalité des larves.相似文献
54.
Even Amler Nelvio Cester Eleonora Salvolini Roberto Staffolani Martin Burkhard Laura Mazzanti Arnot Kotyk Carlo Romanini 《Cell biology international》1994,18(7):723-727
Placentas of women suffering from pregnancy-induced hypertension (PIH) were found to contain a greater amount of Na,K-ATPase molecules, estimated from anthroyl ouabain binding, than normotensive individuals. Both the microsomal fraction of placental cells and purified Na,K-ATPase showed an increased affinity for the specific inhibitor ouabain which, in the case of the microsomes, bound with a dissociation constant of 0.9 nM as compared with 3.4 nM in the controls. Likewise, the dissociation constant of the ouabain complex with purified Na,K-ATPase was about 3.5 times lower in the hypertensive patients. The differences are apparently caused by a different microenvironment of the ouabain-binding site, as reflected in the quantum yield of bound anthroyl ouabain. If an endogenous digitalis-like factor is present in the body fluids to regulate Na,K-ATPase activity, the present results render its role quite plausible. 相似文献
55.
56.
Wing Y. Cheung Jean-Charles Côté Diane L. Benoit Benoit S. Landry 《Plant Molecular Biology Reporter》1993,11(2):142-155
We have designed a simple and rapid assay for chloroplast-based triazine resistance in higher plants using PCR amplification
of thepsbA gene coupled toMaeI digestion of the amplified product to distinguish triazine resistant from sensitive biotypes. Our assay is universal and
avoids the need of lengthy procedures of previously published assays, which either required spraying of seedlings in a controlled
environment, quantification of chlorophyll fluorescence of leaf discs after incubation in triazine solution, DNA sequencing
of thepsbA gene, or Southern-blot analysis. Our diagnostic system is qualitative, reliable, fast and simple. More than 100 seedlings
taken directly from the field can be analyzed in one day. This system has a direct application towards a more rational use
of herbicides in production fields. It also represents a valuable tool to monitor spreading of resistant biotypes through
time and space and can serve as a model system applicable to other gene monitoring needs. 相似文献
57.
Isolation of polysaccharide-free DNA from plants 总被引:2,自引:2,他引:0
Benoît Rether Geneviève Delmas Abdelkrim Laouedj 《Plant Molecular Biology Reporter》1993,11(4):333-337
A quick procedure for the isolation of polysaccharide-free DNA from different plant species and cell suspension or callus
cultures is described. The originality of the method lies in the use of a mixture of glycoside hydrolases that leads, after
phenol and chloroform extraction, to the isolation of pure DNA without any polysaccharide contamination. The highly purified
DNA can be used for nucleotide analysis by HPLC, RFLP analysis and PCR amplification. 相似文献
58.
OBJECTIVES: To determine the prevalence and risk indicators of cervical infection due to Chlamydia trachomatis among female patients consulting for contraception and to evaluate an enzyme immunoassay for the detection of C. trachomatis in this setting. DESIGN: Prevalence study. Endocervical specimens were analysed by means of culture and enzyme immunoassay. C. trachomatis infection was diagnosed through culture. SETTING: A hospital family planning clinic in Trois-Rivières, Que. SUBJECTS: All 533 female patients who consulted for contraception between November 1986 and March 1988. Results of culture were available for 495 patients. MAIN OUTCOME MEASURE: Demographic, epidemiologic and clinical information was collected by means of a standard questionnaire and a gynecologic examination. MAIN RESULTS: The prevalence rate of chlamydial infection was 9% (45/495). Enzyme immunoassay detected 37 (82%) of the infections. The mean age of the patients was 19.8 years, and 98% of the infections were diagnosed in those aged 25 years or less. The variables significantly associated with C. trachomatis infection were having more than one sexual partner in the preceding year (odds ratio [OR] 2.9; 95% confidence limits [CL] 1.7 and 5.0) and having more than one partner in the preceding 3 months (OR 2.3; 95% CL 1.2 and 4.3). These two indicators would have detected 58% and 22% of the infections respectively. CONCLUSIONS: Screening for C. trachomatis infection by means of enzyme immunoassay should be proposed to all female patients aged 25 years or less consulting for contraception in our clinic. Such screening may prove to be an effective preventive measure in other similar clinical settings. 相似文献
59.
Le Maître Anne Guy Franck Merceron Gildas Kostopoulos Dimitris S. 《International journal of primatology》2023,44(1):209-236
International Journal of Primatology - Discoveries in recent decades indicate that the large papionin monkeys Paradolipopithecus and Procynocephalus are key members of the Late Pliocene –... 相似文献
60.
Csaba Moskát Tamás Székely Tibor Kisbenedek Zsolt Karcza István Bártol 《Journal of avian biology》2003,34(1):16-19
We tested the importance of nest cleaning in egg rejection behaviour of the great reed warbler Acrocephalus arundinaceus in a highly parasitised population in which about 64% of nests are parasitised by the common cuckoo Cuculus canorus . Three types of objects of the same weight, texture and colour but with different shapes (dummy cuckoo eggs, sticks and disks) were placed into great reed warbler nests. We investigated the response of hosts in two stages of breeding: pre-incubation when the risk of brood parasitism is high, and during incubation when the risk of parasitism is low. The dummy cuckoo eggs were rejected less often than the other objects in both breeding stages, although we did not find any difference in the frequency of rejection between pre-incubation and incubation. We integrate these results into current views on the evolution of host–parasite interactions, and propose a hierarchical concept to understand egg rejection behaviour: (1) hosts reject all non-egg shaped objects as a general cleaning mechanism; (2) adaptations for the hosts' ability to recognise their own eggs allows them to distinguish these eggs from similar objects and parasitic eggs. 相似文献