全文获取类型
收费全文 | 19687篇 |
免费 | 1324篇 |
国内免费 | 6篇 |
学科分类
生物科学 | 21017篇 |
出版年
2023年 | 132篇 |
2022年 | 118篇 |
2021年 | 220篇 |
2020年 | 267篇 |
2019年 | 244篇 |
2018年 | 603篇 |
2017年 | 515篇 |
2016年 | 791篇 |
2015年 | 1169篇 |
2014年 | 1095篇 |
2013年 | 1505篇 |
2012年 | 1794篇 |
2011年 | 1685篇 |
2010年 | 1004篇 |
2009年 | 795篇 |
2008年 | 1255篇 |
2007年 | 1196篇 |
2006年 | 1155篇 |
2005年 | 997篇 |
2004年 | 1008篇 |
2003年 | 890篇 |
2002年 | 835篇 |
2001年 | 137篇 |
2000年 | 99篇 |
1999年 | 136篇 |
1998年 | 126篇 |
1997年 | 81篇 |
1996年 | 97篇 |
1995年 | 94篇 |
1994年 | 84篇 |
1993年 | 85篇 |
1992年 | 74篇 |
1991年 | 56篇 |
1990年 | 48篇 |
1989年 | 43篇 |
1988年 | 39篇 |
1987年 | 29篇 |
1986年 | 24篇 |
1985年 | 43篇 |
1984年 | 48篇 |
1983年 | 27篇 |
1982年 | 39篇 |
1981年 | 21篇 |
1980年 | 19篇 |
1979年 | 21篇 |
1978年 | 17篇 |
1977年 | 12篇 |
1976年 | 28篇 |
1975年 | 22篇 |
1967年 | 11篇 |
排序方式: 共有10000条查询结果,搜索用时 234 毫秒
51.
Viability measurements of hybridoma cells in suspension cultures 总被引:1,自引:0,他引:1
José M. Coco-Martin Jan W. Oberink Tiny A. M. van der Velden-de Groot E. Coen Beuvery 《Cytotechnology》1992,8(1):57-64
Several methods were applied to determine the viability of hybridoma cells in suspension. These methods include dye inclusion
and exclusion assays such as the classical trypan blue exclusion assay, the propidium iodide (PI) exclusion assay and the
fluorescein diacetate (FDA) inclusion assay. Furthermore, the relation was studied between release of lactate dehydrogenase
(LDH) by hybridoma cells and their viability. Also the ATP content of the cells and cellular heterogeneity as measured with
a flow cytometer were determined in relation to cellular viability.
The dye inclusion and exclusion assays using trypan blue, FDA, PI were shown to be useful methods to determine cellular viability.
With the FDA and PI methods it was possible to obtain additional information about cells which are in a transition state between
viable and non-viable. The viability according to the scatter properties of the cells appears to reflect the overall condition
of the cells, although interpretation of the results is difficult. Measurement of LDH release in the culture fluid or the
cytoplasmic ATP content could not be used as parameters for cell viability. 相似文献
52.
José L. Caballero Eduardo Martinez Francisco Malpartida David A. Hopwood 《Molecular & general genetics : MGG》1991,230(3):401-412
Summary Sequence analysis of the actVA region of the actinorhodin biosynthetic gene cluster of Streptomyces coelicolor revealed a succession of six open reading frames (ORFs), all running in the same direction and extending over 5.32 kb. The protein product of actVA-ORF1 strongly resembles that of another gene, elsewhere in the act cluster (actII-ORF2), which codes for a trans-membrane protein previously implicated in actinorhodin export from the mycelium. This suggests that the two gene products may co-operate in actinorhodin export, perhaps being sufficient for self-protection of the organism against suicide. At least four of the other five ORFs are implicated in the control of the C-6 and C-8 ring-hydroxylation reactions, lacking in actVA mutants, that occur at middle to late stages in the actinorhodin biosynthetic pathway. This conclusion was reached by genetic mapping of actVA mutants to actVA-ORF3 and-ORF5 (and perhaps -ORF4), and by the finding of strong resemblances between the protein products of actVA-ORF2 and -ORF6 and the products of genes of the oxytetracycline or tetracenomycin gene clusters that have been implicated in ring-hydroxylation reactions in the biosynthesis of these other aromatic polyketide antibiotics. 相似文献
53.
CO2 exchange between the atmosphere and soil algal crusts of the Trachypogon savannas of the Orinoco Llanos has been analyzed
using an open gas exchange system. These savannas encompass a wide range of physiognomic types, from herbaceous communities
to savanna woodlands. A maximum CO2 flux of 0.207 mg m-2 s-1 was measured in the crusts of the Guanipa savannas, while in the other examined crusts (0.035–0.105 mg m-2 s-1) the flux was similar to values reported for terrestrial algae. The CO2 flux data were statistically fitted to the photosynthetically active radiation by a logarithmic relationship, and the photosynthetic
efficiencies of the crusts were compared. The activation energy calculated for the CO2 fixation indicates that limitations by diffusion and photochemical processes were excluded in the Guanipa crusts (above 12
kcal mole-1), whereas they were evident in the other crust studied. An optimum CO2 incorporation as a function of the crust water potential was established and carbon gain strategies were proposed on the
basis of the results and characteristics of the habitats. 相似文献
54.
The primary structure of the HLA-A2 subtype A*0204 (isoelectric focusing variant A2.A) has been determined. cDNA encoding this subtype was amplified by the polymerase chain reaction. Four independent full-lenght cDNA clones encoding A*0204 were analyzed to obtain a consensus sequence for this subtype. A*0204 differs from A*0201 by a single nucleotide change of G to T through the coding regions, resulting in an Arg to Met change at position 97. This substitution accounts for the isoelectric focusing pattern of the subtype. The same change occurs in other HLA-A specificities in association with other changes in its vicinity. The absence of additional substitutions in A*0204 suggests that it could have arisen from A*0201 by point mutation, and that recurrent mutations may take place during HLA diversification. The spatial location of this change implies that A*0204 must be a functional variant. Comparison of its sequence with other HLA-A2 subtypes reveals that much of the HLA-A2 subtype polymorphism is generated by variations in four neighboring positions, including position 97, which are located in two adjacent -strands on the floor of the peptide binding site of the molecule.The nucleotide sequence data reported in this paper have been submitted to the EMBL nucleotide sequence database and have been assigned the accession number X57954.
Address correspondence and offprint requests to: J. A. López de Castro. 相似文献
55.
Cecilia Zazueta José A. Holguín Jorge Ramírez 《Journal of bioenergetics and biomembranes》1991,23(6):889-902
We describe a calcium transport that is sensitive to ruthenium red in liposomes reconstituted with mitochondrial extracts. This system is able to build an internally negative membrane potential, which allows the electrogenic influx of Ca2+ and Sr2+. Proteins with molecular weights higher than 35 kDa were incorporated to the vesicles, and enhanced the accumulation of the cation in an energy-dependent fashion. 相似文献
56.
Chloramphenicol acetyl transferase (CAT) gene was used as a reporter gene to assess the conditions for polyethylene glycol (PEG)-mediated transfection of kiwifruit protoplasts. The effect of plasmid concentration and the presence of carrier DNA were each assessed by analysing CAT activity in transfected protoplasts using thin-layer chromatography (TLC) autoradiographic detection of acetylated chloramphenicol. A gas chromatography (GC) and gas chromatography-mass spectrometry (GC-MS) non-radioactive method was developed for monitoring CAT gene activity. This method provides a high speed of analysis (30 min) and precise means of detecting acetylated products at the nanomolar level, enabling quantification at very low transfection rates. Using this method we optimized plasmid and PEG concentration and also assessed the effect of heat shock on transfection. The best CAT activity was obtained using 30% polyethylene glycol 4000 and by submitting protoplasts to heat shock (45 °C, 5 min) prior to transfection. 相似文献
57.
Interaction of two variable proteins (PilE and PilC) required for pilus-mediated adherence of Neisseria gonorrhoeae to human epithelial cells 总被引:16,自引:0,他引:16
Thomas Rudel Jos P. M. van Putten Carol P. Gibbs † Rainer Haas Thomas F. Meyer 《Molecular microbiology》1992,6(22):3439-3450
Pili confer the initial ability of Neisseria gonorrhoeae to bind to epithelial cells. Pilin (PilE), the major pilus subunit, and a minor protein termed PilC, reportedly essential for pilus biogenesis, undergo intra-strain phase and structural variation. We demonstrate here that at least two different adherence properties are associated with the gonococcal pili: one is specific for erythrocytes, which is virtually unaffected by PilE variation, and another is specific for epithelial cells, and is modulated in response to the variation of PilE. Based on this finding, mutants of a recA- strain were selected that had lost the ability to bind to human cornea epithelial cells (A-) but retained the ability to form pili (P+) and to agglutinate human erythrocytes (H+). The adherence-negative mutants failed to produce detectable levels of PilC1 or PilC2 proteins, representing piIC phase variants generated in the absence of RecA. The A- pilC phase variants were indistinguishable from their A+ parents and spontaneous A+ revertants with regard to the amount of PilE produced and its electrophoretic mobility, the degrees of piliation and haemagglutination, and the pilE nucleotide sequence. These data demonstrate a central role for PilC in pilus-mediated adherence of N. gonorrhoeae to human epithelial cells and further indicate that neither PilC1 nor PilC2 is obligatory for the assembly of gonococcal pili. 相似文献
58.
Arturo Manjón José L. Iborra Concepción Martínez-Madrid 《Applied microbiology and biotechnology》1991,35(2):176-179
Summary
Rhodococcus fascians cells were immobilized by entrapment in -carrageenan. The ability of the system to continuously degrade limonin was tested against pH. A burst of activity was observed when changing from pH 4.5 to 5.0, and a small increase could be seen above the latter value. Such behaviour was not only a response of the metabolic activity of the cells to changes in the medium pH, but to selectivity towards the chemical form of the limonin substrate, which also depends on pH. Additionally, the immobilized cells showed increased resistance against pH changes, since the system recovered almost full activity when the pH was restored to 7.0 after being operated for long periods at pH 4.0. The decrease in limonin-degrading capability of the immobilized cells at low pH values could be overcome by choosing an appropriate dilution rate.Offprint requests to: J. L. Iborra 相似文献
59.
Pigment patterns in mutants affecting the biosynthesis of pteridines and xanthommatin in Drosophila melanogaster 总被引:3,自引:0,他引:3
Juan Ferré Francisco J. Silva M. Dolores Real José L. Ménsua 《Biochemical genetics》1986,24(7-8):545-569
Eye-color mutants of Drosophila melanogaster have been analyzed for their pigment content and related metabolites. Xanthommatin and dihydroxanthommatin (pigments causing brown eye color) were measured after selective extraction in acidified butanol. Pteridines (pigments causing red eye color) were quantitated after separation of 28 spots by thin-layer chromatography, most of which are pteridines and a few of which are fluorescent metabolites from the xanthommatin pathway. Pigment patterns have been studied in 45 loci. The pteridine pathway ramifies into two double branches giving rise to isoxanthopterin, drosopterins, and biopterin as final products. The regulatory relationship among the branches and the metabolic blockage of the mutants are discussed. The Hn locus is proposed to regulate pteridine synthesis in a step between pyruvoyltetrahydropterin and dihydropterin. The results also indicate that the synthesis and accumulation of xanthommatin in the eyes might be related to the synthesis of pteridines.Support for this work was provided to J.F. in part by a grant from the Ministerio de Universidades e Investigación (Spain) and to F.J.S. by a grant from the Ministerio de Educación y Ciencia (Spain). 相似文献
60.