全文获取类型
收费全文 | 424723篇 |
免费 | 47913篇 |
国内免费 | 162篇 |
出版年
2018年 | 3903篇 |
2016年 | 5284篇 |
2015年 | 6904篇 |
2014年 | 8150篇 |
2013年 | 11240篇 |
2012年 | 12786篇 |
2011年 | 13235篇 |
2010年 | 9074篇 |
2009年 | 8427篇 |
2008年 | 12113篇 |
2007年 | 12611篇 |
2006年 | 11825篇 |
2005年 | 11300篇 |
2004年 | 11343篇 |
2003年 | 10631篇 |
2002年 | 10453篇 |
2001年 | 17418篇 |
2000年 | 17428篇 |
1999年 | 13910篇 |
1998年 | 5020篇 |
1997年 | 5270篇 |
1996年 | 4911篇 |
1995年 | 4616篇 |
1994年 | 4483篇 |
1993年 | 4536篇 |
1992年 | 11607篇 |
1991年 | 11534篇 |
1990年 | 11292篇 |
1989年 | 10909篇 |
1988年 | 10486篇 |
1987年 | 10075篇 |
1986年 | 9363篇 |
1985年 | 9233篇 |
1984年 | 7749篇 |
1983年 | 6703篇 |
1982年 | 5162篇 |
1981年 | 4616篇 |
1980年 | 4463篇 |
1979年 | 7420篇 |
1978年 | 5863篇 |
1977年 | 5398篇 |
1976年 | 5203篇 |
1975年 | 5596篇 |
1974年 | 6302篇 |
1973年 | 6166篇 |
1972年 | 5766篇 |
1971年 | 5226篇 |
1970年 | 4634篇 |
1969年 | 4575篇 |
1968年 | 4422篇 |
排序方式: 共有10000条查询结果,搜索用时 15 毫秒
211.
Monitoring signal transduction in cancer: cDNA microarray for semiquantitative analysis. 总被引:1,自引:0,他引:1
H B Hsieh R A Lersch D E Callahan S Hayward M Wong O H Clark H U Weier 《The journal of histochemistry and cytochemistry》2001,49(8):1057-1058
This study targeted the development of a novel microarray tool to allow rapid determination of the expression levels of 58 different tyrosine kinase (tk) genes in small tumor samples. The goals were to define a reference probe for multi-sample comparison and to investigate the variability and reproducibility of the image acquisition and RT-PCR procedures. The small number of tk genes on our arrays enabled us to define a reference probe by artificially mixing all genes on the arrays. Such a probe provided contrast reference for comparative hybridization of control and sample DNA and enabled cross-comparison of more than two samples against one another. Comparison of signals generated from multiple scanning eliminated the concern of photo bleaching and scanner intrinsic noise. Tests performed with breast, thyroid, and prostate cancer samples yielded distinctive patterns and suggest the feasibility of our approach. Repeated experiments indicated reproducibility of such arrays. Up- or downregulated genes identified by this rapid screening are now being investigated with techniques such as in situ hybridization. 相似文献
212.
Concentrations of the elements C, N, Fe, Na, K, Ca and Mg were analysed by chromatography and atomic absorption spectrometry in the whaleworm Anisakis simplex sensu lato. Nearly all mean inorganic element concentrations measured were higher in adult worms than in larval forms. Similarly, adults contained significantly higher nitrogen (i.e. lower C/N ratio values) than either eggs or larvae. Evidence for the significance of the parasitic life cycle stage and the systematic position of their hosts on the element concentration variability is discussed. 相似文献
213.
G. I. Novik N. I. Astapovich Zh. N. Bogdanovskaya N. E. Ryabaya 《Applied Biochemistry and Microbiology》2000,36(1):90-95
Production of Enterobifidin includes the stages of preparation of culture media, reparation of lyophilizedBifidobacterium adolescentis MS-42 culture, preparation of starters, cultivation of bacteria in fermenters, biomass conservation, and its biological control.
The preparation contains physiologically active bifidobacterium cells with high activities of growth(μ = 0.7 h−1,g = 1.0 h) and acid formation (titratable acidity is ∼120–140°T; acetate concentration, 0.50–0.75%; and lactate concentration,
0.33–0.50%). The antagonistic activity of these bacteria towardsEscherichia coli 08,E. coli 086,E. coli 015,E. coli 0115, andE. coli 0101 amounts to 98.2; toProteus vulgaris 102, to 87.2; andStaphylococcus aureus 209p, to 83.2%. The bifidobacteria (with a titer of ∼109 CFU/ml) remained viable for two to five months. 相似文献
214.
215.
216.
Optimized linker sequences for the expression of monomeric and dimeric bispecific single-chain diabodies. 总被引:5,自引:0,他引:5
Bispecific single-chain diabodies (scDb) consist of the variable heavy and light chain domains of two antibodies connected by three linkers. The structure of an scDb in the V(H)-V(L) orientation is V(H)A-linkerA-V(L)B-linkerM-V(H)B-linkerB-V(L)A, with linkers A and B routinely chosen to be 5-6 residues and linker M 15-20 residues. Here, we applied display of scDb on filamentous phage to analyse the composition of optimal linker sequences. The three linkers were randomized in length and sequence using degenerated triplets coding for only six hydrophilic or aliphatic amino acids (Thr, Ser, Asp, Asn, Gly, Ala). Antigen-binding clones were then isolated by one to two rounds of selection on the two different antigens recognized by the bispecific scDb. Using an scDb directed against carcinoembryonic antigen (CEA) and beta-galactosidase (Gal), we found that monomeric scDb had a preferred length of 15 or more amino acid residues for the middle linker M and of 3-6 residues for the linkers A and B. No obvious bias towards a preferred linker sequence was observed. Reduction of the middle linker below 13 residues led to the formation of dimeric scDb, which most likely results from interchain pairing between all the V(H) and V(L) domains. Dimeric scDb were also formed by fragments possessing a long linker M and linkers A and B of 0 or 1 residue. We assume that these dimeric scDb are formed by intrachain pairing of the central variable domains and interchain pairing of the flanking variable domains. Thus, the latter molecules represent a novel format of bispecific and tetravalent molecules. The described strategy allows for the isolation of both optimized and minimal linker sequences for the assembly of monomeric or dimeric single-chain diabodies. 相似文献
217.
Shira Weingarten-Gabbay Susan Klaeger Siranush Sarkizova Leah R. Pearlman Da-Yuan Chen Kathleen M.E. Gallagher Matthew R. Bauer Hannah B. Taylor W. Augustine Dunn Christina Tarr John Sidney Suzanna Rachimi Hasahn L. Conway Katelin Katsis Yuntong Wang Del Leistritz-Edwards Melissa R. Durkin Christopher H. Tomkins-Tinch Pardis C. Sabeti 《Cell》2021,184(15):3962-3980.e17
218.
We re-engineered a classic tool for mutagenesis and gene expression studies in Gram-negative bacteria. Our modified Tn5-based transposon contains multiple features that allow rapid selection for mutants, direct quantification of gene expression and straightforward cloning of the inactivated gene. The promoter-less gfp-km cassette provides selection and reporter assay depending on the activity of the promoter upstream of the transposon insertion site. The cat gene facilitates positive antibiotic selection for mutants, while the narrow R6Kγ replication origin forces transposition in recipient strains lacking the pir gene and enables cloning of the transposon flanked with the disrupted gene from the chromosome. The suicide vector pCKD100, a plasmid that could be delivered into recipient cells through biparental mating or electroporation, harbours the modified transposon. We used the transposon to mutagenize Pectobacterium versatile KD100, Pseudumonas coronafaciens PC27R and Escherichia coli 35150N. The fluorescence intensities of mutants expressing high GFP could be quantified and detected qualitatively. Transformation efficiency from conjugation ranged from 1600 to 1900 CFU per ml. We sequenced the upstream flanking regions, identified the putative truncated genes and demonstrated the restoration of the GFP phenotype through marker exchange. The mini-Tn5 transposon was also utilized to construct mutant a library of P. versatile for forward genetic screens. 相似文献
219.
D.A.M. da Silva M.S. Fernandes E.H. Endo A.C.P. Vital E.A. Britta M.E. Favero J.C. Castro P.T. Matumoto-Pintro B.P. Dias Filho C.V. Nakamura M. Machinski Junior J.M.G. Mikcha B.A. de Abreu Filho 《Letters in applied microbiology》2021,72(1):41-52
The use of rosemary essential oil (RO) and its combination with nisin (RO+N) in preventing the multiplication of Alicyclobacillus acidoterrestris in orange juice was evaluated. The minimum inhibitory and bactericidal concentrations (MIC and MBC) for RO were both 125 μg ml−1 while RO+N displayed a synergistic effect. The use of RO and RO+N at concentrations of 1, 4 and 8× MIC in orange juice for 96 h was evaluated in terms of their sporicidal effectiveness. With regard to the action against A. acidoterrestris spores, RO at 8× MIC was sporostatic, whereas RO+N at 1× MIC was sporicidal. Morphological changes in the structure of the micro-organism after treatment were also observed by microscopy. Furthermore, flow cytometric analysis showed that most cells were damaged or killed after treatment. In general, the antioxidant activity after addition of RO+N decreased with time. The results demonstrate that using the combination of RO and nisin can prevent the A. acidoterrestris growth in orange juice. 相似文献
220.
F. Hubert M.-H. Rodier A. Minoza V. Portet-Sulla E. Cateau K. Brunet 《Letters in applied microbiology》2021,72(1):82-89
Candida auris is an emerging species responsible for life-threatening infections. Its ability to be resistant to most systemic antifungal classes and its capacity to persist in a hospital environment have led to health concerns. Currently, data about environmental reservoirs are limited but remain essential in control of C. auris spread. The aim of our study was to explore the interactions between C. auris and two free-living amoeba (FLA) species, Vermamoeba vermiformis and Acanthamoeba castellanii, potentially found in the same water environment. Candida auris was incubated with FLA trophozoites or their culture supernatants. The number of FLA and yeasts was determined at different times and transmission electron microscopy (TEM) was performed. Supernatants of FLAs promoted yeast survival and proliferation. Internalization of viable C. auris within both FLA species was also evidenced by TEM. A water environmental reservoir of C. auris can therefore be considered through FLAs and contamination of the hospital water networks would consequently be possible. 相似文献