全文获取类型
收费全文 | 3383篇 |
免费 | 199篇 |
国内免费 | 9篇 |
出版年
2023年 | 49篇 |
2022年 | 125篇 |
2021年 | 158篇 |
2020年 | 97篇 |
2019年 | 124篇 |
2018年 | 125篇 |
2017年 | 85篇 |
2016年 | 133篇 |
2015年 | 168篇 |
2014年 | 189篇 |
2013年 | 275篇 |
2012年 | 228篇 |
2011年 | 269篇 |
2010年 | 135篇 |
2009年 | 130篇 |
2008年 | 133篇 |
2007年 | 149篇 |
2006年 | 124篇 |
2005年 | 109篇 |
2004年 | 102篇 |
2003年 | 86篇 |
2002年 | 90篇 |
2001年 | 48篇 |
2000年 | 30篇 |
1999年 | 38篇 |
1998年 | 25篇 |
1997年 | 23篇 |
1996年 | 13篇 |
1995年 | 11篇 |
1994年 | 22篇 |
1993年 | 13篇 |
1992年 | 17篇 |
1991年 | 17篇 |
1990年 | 13篇 |
1989年 | 14篇 |
1988年 | 24篇 |
1987年 | 13篇 |
1986年 | 18篇 |
1985年 | 17篇 |
1984年 | 9篇 |
1983年 | 14篇 |
1982年 | 12篇 |
1981年 | 8篇 |
1980年 | 8篇 |
1979年 | 13篇 |
1977年 | 12篇 |
1976年 | 10篇 |
1974年 | 6篇 |
1973年 | 10篇 |
1971年 | 7篇 |
排序方式: 共有3591条查询结果,搜索用时 31 毫秒
961.
Genetics in the age of systems biology 总被引:3,自引:0,他引:3
Systems biology has become a fashionable label for a new generation of large-scale experiments. This essay explores how classical approaches such as forward genetics fit into this emerging framework. 相似文献
962.
963.
Jeon WB Aceti DJ Bingman CA Vojtik FC Olson AC Ellefson JM McCombs JE Sreenath HK Blommel PG Seder KD Burns BT Geetha HV Harms AC Sabat G Sussman MR Fox BG Phillips GN 《Journal of structural and functional genomics》2005,6(2-3):143-147
The Center for Eukaryotic Structural Genomics (CESG) has established procedures for the purification of Arabidopsis proteins in a high-throughput mode. Recombinant proteins were fused with (His)(6)-MBP tags at their N-terminus and expressed in Escherichia coli. Using an automated AKTApurifier system, fusion proteins were initially purified by immobilized metal affinity chromatography (IMAC). After cleavage of (His)(6)-MBP tags by TEV protease, (His)(6)-MBP tags were separated from target proteins by a subtractive 2nd IMAC. As a part of quality assurance, all purified proteins were subjected to MALDI-TOF and ESI mass spectrometry to confirm target identity and integrity, and determine incorporation of seleno-methionine (SeMet) and (15)N and (13)C isotopes. The protocols have been used successfully to provide high quality proteins that are suitable for structural studies by X-ray crystallography and NMR. 相似文献
964.
Schmitz M Alfalah M Aerts JM Naim HY Zimmer KP 《The international journal of biochemistry & cell biology》2005,37(11):2310-2320
Gaucher's disease is the most inherited lysosomal storage disorder. Except for a few cases, the broad phenotypic heterogeneity of Gaucher's disease can be neither predicted from defined mutations nor from differences in residual enzyme activity. Here, we analyse the intracellular trafficking of glucocerebrosidase as an underlying mechanism for the expression of the clinical phenotype. Biosynthetic labeling studies combined with immunofluorescence analyses with fibroblasts from patients with the defined mutations N370S, L444P, D409H and G202R unequivocally demonstrate a retarded transport of glucocerebrosidase carrying the mutation N370S and a transport block in the ER of the enzyme with the mutations G202R, L444P and D409H. We asked whether cellular components in the patients' fibroblasts other than glucocerebrosidase are implicated in the onset of the disease. For this, mutant cDNA's corresponding to the phenotypes N370S, G202R and L444P were expressed in the mouse fibroblasts NIH3T3. Essentially similar biochemical and cellular features were revealed as compared to the patients' fibroblasts strongly suggesting that these mutations are exclusively responsible for the characterized phenotypes. Interestingly, the immunoglobulin binding protein (BiP) binds wild type and the mutant N370S but not the G202R and L444P variants suggesting a discriminatory role played by this chaperone associated with the severity of the disease. 相似文献
965.
Fathallah-Shaykh HM 《Bioinformatics (Oxford, England)》2005,21(23):4255-4262
SUMMARY: MASH is a mathematical algorithm that discovers highly specific states of expression from genomic profiling by microarrays. The goal at the outset of this analysis was to improve the sensitivity of MASH. The geometrical representations of microarray datasets in the 3D space are rank-dependent and unique to each dataset. The first filter (F1) of MASH defines a zone of instability whose F1-sensitive ratios have large variations. A new filter (Fs) constructs in the 3D space rank-dependent lower and upper-bound contour surfaces, which are modeled based on the geometry of the unique noise intrinsic to each dataset. As compared with MASH, Fs increases sensitivity significantly without lowering the high specificity of discovery. Fs facilitates studies in functional genomics and systems biology. 相似文献
966.
967.
Tyler RC Sreenath HK Singh S Aceti DJ Bingman CA Markley JL Fox BG 《Protein expression and purification》2005,40(2):268-278
Protocols have been developed and applied for the high-throughput production of [U-15N]- or [U-13C-, U-15N]-labeled proteins using the conditional methionine auxotroph Escherichia coli B834. The large-scale growth and expression uses a chemically defined auto-induction medium containing salts and trace metals, vitamins including vitamin B12, and glucose, glycerol, and lactose. The results from nine expression trials in 2-L of the auto-induction medium (500 mL in each of four polyethylene terephthalate beverage bottles) gave an average final optical density at 600 nm of approximately 5, an average wet cell mass yield of approximately 9.5 g L(-1), and an average yield of approximately 20 mg of labeled protein in the six instances in which proteolysis of the fusion protein was observed. Correlations between the cell mass recovered, the level of protein expression, and the relative amounts of glucose, glycerol, and lactose in the auto-induction medium were noted. Mass spectral analysis showed that the purified proteins contained both 15N and 13C at levels greater than 95%. 1H-15N heteronuclear single quantum correlation spectroscopy as well as 13C; 15N-edited spectroscopy showed that the purified [U-15N]- and [U-13C, U-15N]-labeled proteins were suitable for structure analysis. 相似文献
968.
Sreenath HK Bingman CA Buchan BW Seder KD Burns BT Geetha HV Jeon WB Vojtik FC Aceti DJ Frederick RO Phillips GN Fox BG 《Protein expression and purification》2005,40(2):256-267
Protocols have been developed and applied in the high-throughput production of selenomethionine labeled fusion proteins using the conditional Met auxotroph Escherichia coli B834. The large-scale growth and expression uses a chemically defined auto-induction medium containing 125 mg L(-1) selenomethionine, salts and trace metals, other amino acids including 10 mg L(-1) of methionine, vitamins except vitamin B12, and glucose, glycerol, and alpha-lactose. A schematic for a shaker rack that can hold up to twenty-four 2-L polyethylene terephthalate beverage bottles in a standard laboratory refrigerated floor shaker is provided. The growth cycle from inoculation of the culture bottle through the growth, induction, and expression was timed to take approximately 24 h. Culture growth in the auto-induction medium gave an average final optical density at 600 nm of approximately 6 and an average wet cell mass yield of approximately 14 g from 2 L of culture in greater than 150 expression trials. A simple method for visual scoring of denaturing electrophoresis gels for total protein expression, solubility, and effectiveness of fusion protein proteolysis was developed and applied. For the favorably scored expression trials, the average yield of purified, selenomethionine-labeled target protein obtained after proteolysis of the fusion protein was approximately 30 mg. Analysis by mass spectrometry showed greater than 90% incorporation of selenomethionine over a approximately 8-fold range of selenomethionine concentrations in the growth medium, with higher growth rates observed at the lower selenomethionine concentrations. These protein preparations have been utilized to solve X-ray crystal structures by multiwavelength anomalous diffraction phasing. 相似文献
969.
Khan IH Kim H Ashida H Ishikawa T Shibata H Sawa Y 《Bioscience, biotechnology, and biochemistry》2005,69(9):1802-1805
The Lys80, Gly82 and Met101 residues of glutamate dehydrogenase from Bacillus subtilis were mutated into a series of single mutants. The wild-type enzyme was highly specific for 2-oxoglutarate, whereas G82K and M101S dramatically switched to increased specificity for oxaloacetate with kcat values 3.45 and 5.68 s-1, which were 265-fold and 473-fold higher respectively than those for 2-oxoglutarate. 相似文献
970.
Application of lipases in kinetic resolution of racemates 总被引:3,自引:0,他引:3
Lipases have been well established as valuable catalysts in organic synthesis. This review article focuses on some of the recent developments in the rapidly growing field of lipase-catalyzed kinetic resolution of racemates as a versatile method for the separation of enantiomers. The literature search dates back to the last five years and covers some comprehensive examples. The main emphasis is on the use of lipases in organic solvents. 相似文献