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11.
The erythromycin resistance plasmid pSM752 carrying the cloned streptokinase gene, skc, was introduced by protoplast transformation into Streptococcus equisimilis H46A from which skc was originally cloned. Cells transiently supporting the replication of pSM752 gave rise to an erythromycin-resistant clone designated H46SM which was plasmid free and produced streptokinase at levels approximately twice as high as the wild type. Southern hybridization of total cell DNA with an skc-containing probe provided evidence for the duplication of the skc gene in the H46SM chromosome. The results, which have some bearing on industrial streptokinase production, can be best explained by a single cross-over event between the chromosome and the plasmid in the region of shared homology leading to the integration of pSM752 in a Campbell-like manner. 相似文献
12.
Tjakko Abee Jan Knol Klaas J. Hellingwerf Evert P. Bakker Annette Siebers Wil N. Konings 《Archives of microbiology》1992,158(5):374-380
Cells of the purple non-sulphur bacterium Rhodobacter sphaeroides express a high-affinity K+ uptake system when grown in media with low K+ concentrations. Antibodies againts the catalytic KdpB protein or the whole KdpABC complex of Escherichia coli crossreact with a 70.0 kDa R. sphaeroides protein that was expressed only in cells grown in media with low K+ concentrations. In membranes derived from R. sphaeroides cells grown with low K+ concentrations (induced cells), a high ATPase activity could be detected when assayed in Tris-HCl pH 8.0 containing 1 mM MgSO4. This ATPase activity increased upon addition of 1 mM KCl from 166 to 289 mol ATP hydrolysed x min-1 x g protein-1 (1.7-fold stimulation). The K+-stimulated ATPase activity was inhibited approximately 93% by 0.5 mM vanadate but hardly by N,N-dicyclohexylcarbo-diimide (DCCD). These results indicate that the inducible K+-ATPase in R. sphaeroides resembles the Kdp K+-translocating ATPase of Escherichia coli. This Kdp-like transport system is also expressed in R. capsulatus and Rhodospirillum rubrum during growth in media with low K+ concentrations suggesting a wide distribution of this transport system among phototrophic bacteria.Abbreviations
electrical potential difference across the cytoplasmic membrane
- pH
pH difference across the cytoplasmic membrane
- BSA
bovine serum albumine
- PAGE
polyacrylamide gel electrophoresis
- HEPES
4-(2-hydroxyethyl)-1-piperazine-ethanesulfonic acid
- PMSF
phenyl-methyl-sulfonyl fluoride
- DCCD
N,N-dicyclohexylcarbodiimide
- AIB
2--aminoisobutyric acid
- TMG
methyl--d-thiogalactopyranoside 相似文献
13.
Nacyra Assad-García Neftali Ochoa-Alejo Enrique García-Hernández Luis Herrera-Estrella June Simpson 《Plant cell reports》1992,11(11):558-562
A protocol for the Agrobacterium-mediated transformation of tomatillo was developed. Up to 40 transgenic plants could be obtained in experiments using 60 cotyledon expiants. The transformed nature of the regenerated plants was confirmed by NPT II and Southern blot hybridization analysis. Using the b-glucuronidase system the tissue specific and developmental patterns of expression of the Cauliflower Mosaic Virus 35S promoter were determined in transgenic tomatillo plants. It was found that this promoter is developmentally regulated during fruit and seed formation. 相似文献
14.
D. C. Spaulding D. E. Morse 《Journal of comparative physiology. B, Biochemical, systemic, and environmental physiology》1991,161(5):498-515
Summary The digestive glands of many marine molluscs are rich sources of arylsufatase enzymes which may function in the catabolism of sulfated polysaccharides in the diets of herbivorous species. Arylsulfatases, partially purified from the hepatopancreas of the red abalone, Haliotis rufescens, were investigated with respect to heterogeneity, catalytic requirements, and timing of induction during development. Four hepatopancreatic enzymes were purified from adult animals using a combination of hydrophobic interaction and anion-exchange chromatography. Zymograms of the four partially-purified enzymes produced by electrophoresis under nondenaturing conditions revealed a fifth, relatively more basic isozyme. All four partially-purified enzymes appear to be monomeric, with molecular weights of approximately 43 000 Da each, as measured by gel filtration. The affinities for p-nitrocatechol sulfate, pH optima, and strengths of inhibition by anions displayed by these enzymes are similar to the values reported for other molluscan arylsulfatases. Three of the four enzymes have K
m values between 0.8 and 2.0 mM for p-nitrocatechol sulfate; the remaining enzyme (A2) has a K
m of 6.7 mM. All four enzymes have pH and temperature optima of 5.5 and 45°C, respectively. Three of the four enzymes have-t1/2(50°C) values of 3.5 min; the enzyme A4 has a t1/2 has a t1/2(50°C) of 8.5 min. A monoclonal antibody directed against form A1b does not cross react with any of the other hepatopancreatic arylsulfatases when assayed by Western blot, confirming the structural heterogeneity of the adult enzymes.Total arylsulfatase activity increases in a biphasic manner during early abalone development, with the first increase occurring early in larval maturation. The secoad phase of enzyme expression is dependent upon the induction of settlement and metamorphosis of the competent veliger larvae, strongly suggesting that the expression of arylsulfatase synthesis (and the maturation of the digestive gland, the hepatopancreas) is controlled by genetic events which occur as a result of metamorphosis. Competent veliger larvae express only two arylsulfatase forms, which share many physicochemical and kinetic characteristics with the adult hepatopancreatic enzymes. However, neither of the larval arylsulfatases is recognized by the monoclonal antibody to form A1b from adult hepatopancreas. Endogenous enzyme inhibitor levels in larvae remain constant throughout the period of arylsulfatase induction, and therefore do not contribute to the control of arylsulfatase activity levels during development.These results are the first documentation of the developmental induction of a specific protein(s) in abalone as a result of metamorphosis. The significance of the timing of arylsulfatase expression is discussed in relation to potential physiological substrates and the dietary switching which occurs at metamorphosis. Possible genetic events which are consistent with the observed patterns of expression of these enzymes also are considered.Abbreviations
BSA
bovine serum albumin
-
C
centigrade
-
Da
daltons
-
DEAE
diethylaminoethyl
-
ELISA
enzyme-linked immunosorbent assay
-
FPLC
fast protein and polynucleotide liquid chromatography
-
GABA
-aminobutyric acid
- HAT
hypoxanthine, aminopterin, thymidine
- Hepes
N-(2-Hydroxythyl)piperazine-N'-(2-ethanesulfonic acid)
-
HPLC
high pressure liquid chromatography
-
KBS
Kantor's balanced salt solution
-
K
m
Michaelis constant
-
PBS
phosphate buffered saline
-
R
m
relative mobility
-
RPMI
Roswell Park Memorial Institute
-
SDS
sodium dodecyl sulfate
-
T
time
-
TBS
TRIS buffered saline
-
V
max
maximal velocity 相似文献
15.
16.
17.
肝再生的调控及原癌基因表达 总被引:4,自引:0,他引:4
肝脏再生过程中受到多种体液因素的调控。肝细胞生长因子(HGF)、肝刺激因子(HSS)等对肝细胞有促分裂作用;转化生长因子β1(TGFβ1)等则具有抑制作用。此外,肝再生还需要去甲肾上腺素(NE)、胰岛素等辅助分裂原的存在,共同调节肝再生。肝细胞分裂增殖与原癌基因表达密切相关。肝细胞从静息期进入细胞周期,以及在整个细胞周期中,某些原癌基因有特征性的表达。 相似文献
18.
Satoshi Iuchi Kazuko Yamaguchi-Shinozaki Takeshi Urao Kazuo Shinozaki 《Journal of plant research》1996,109(4):415-424
Ten cDNAs for drought-inducible genes were isolated using differential screening of a cDNA library prepared from 10-hr dehydrated
cowpea plants,Vigna unguiculata (S. Iuchi, K. Yamaguchi-Shinozaki, T. Urao, T. Terao, K. Shinozaki; Plant Cell Physiology, 1996 in press). Two of the cDNA
clones, designated CPRD12 and CPRD46, were sequenced and characterized. The CPRD12 and CPRD46 cDNAs encode putative proteins
related to nonmetallo-short-chain alcohol dehydrogenase (CPRD12) and chloroplastic lipoxygenase (CPRD46). Northern blot analysis
revealed that these genes are induced by high-salinity stress and exogenous abscisic acid, but not by cold stress. The CPRD46
gene is also responsive to heat stress and methyl jasmonate and salicylic acid. Genomic Southern blot analysis suggested that
CPRD12 constitutes a small gene family, but that CPRD46 is a single copy gene. We discuss the possible functions of these
two CPRD gene products under drought stress. 相似文献
19.
在自生异养条件下,旋扭山绿豆根瘤菌MXDI6菌株的氢酶诱导表达受气相、pH值、镍等因子影响:氢酶表达的最适氧浓度为4%,最适氢浓度为15%,二氧化碳没有明显影响;氢酶表达的pH值以5.0—6.0为宜;0.5μmol/LNiCl2明显促进吸氢活性,但镍浓度大于1μmol/L则抑制吸氢活性. 相似文献
20.
利用人粒细胞集落刺激因子(hG-CSF)cDNA3′端非翻译区(3′-UTR)中存在的DraⅠ酶切位点,通过部分酶切与完全酶切,删除3′-UTR不同长度,构建了四种hG-CSFcDNA瞬时重组表达质粒。转染COS-7细胞后,生物活性测定结果提示,hG-CSFcDNA3′-UTR对其表达起负调控作用,其关键性序列位于紧接终止密码子TGA下游的65bp范围内,3′-UTR对hG-CSFcDNA表达的影响与转录水平的差别有一定关系。 相似文献