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941.
The extent of the change in thermal diffusivity of soft tissues due to heat-induced damage is not well known. Reported here are the results of using the flash method to measure the through-the-wall component of thermal diffusivity of bovine aorta before and after the tissue has undergone two hours of heating at 75 degrees C. The measurements indicate a 10.1 percent increase in the thermal diffusivity of the tissue post-heating. While this change may not result in a significant change in the tissue temperature profile, further study is needed to quantify the thermal diffusivity in other coordinate directions, as well as the mechanisms by which this change in properties occurs.  相似文献   
942.
943.
The Arbitrary Indian: The Indian Arts and Crafts Act of 1990. Gail K. Sheffield. Norman: University of Oklahoma Press, 1997. 223 pp.  相似文献   
944.
Whole-cell assays of methane and trichloroethylene (TCE) consumption have been performed on Methylosinus trichosporium OB3b expressing particulate methane monooxygenase (pMMO). From these assays it is apparent that varying the growth concentration of copper causes a change in the kinetics of methane and TCE degradation. For M. trichosporium OB3b, increasing the copper growth concentration from 2.5 to 20 μM caused the maximal degradation rate of methane (Vmax) to decrease from 300 to 82 nmol of methane/min/mg of protein. The methane concentration at half the maximal degradation rate (Ks) also decreased from 62 to 8.3 μM. The pseudo-first-order rate constant for methane, Vmax/Ks, doubled from 4.9 × 10−3 to 9.9 × 10−3 liters/min/mg of protein, however, as the growth concentration of copper increased from 2.5 to 20 μM. TCE degradation by M. trichosporium OB3b was also examined with varying copper and formate concentrations. M. trichosporium OB3b grown with 2.5 μM copper was unable to degrade TCE in both the absence and presence of an exogenous source of reducing equivalents in the form of formate. Cells grown with 20 μM copper, however, were able to degrade TCE regardless of whether formate was provided. Without formate the Vmax for TCE was 2.5 nmol/min/mg of protein, while providing formate increased the Vmax to 4.1 nmol/min/mg of protein. The affinity for TCE also increased with increasing copper, as seen by a change in Ks from 36 to 7.9 μM. Vmax/Ks for TCE degradation by pMMO also increased from 6.9 × 10−5 to 5.2 × 10−4 liters/min/mg of protein with the addition of formate. From these whole-cell studies it is apparent that the amount of copper available is critical in determining the oxidation of substrates in methanotrophs that are expressing only pMMO.  相似文献   
945.
16-Fluoropalmitic acid was synthesized from 16-hydroxypalmitic acid using diethylaminosulfur trifluoride. This monofluorinated fatty acid then was used to make 1-palmitoyl-2-[16-fluoropalmitoyl]-phosphatidylcholine (F-DPPC) as a fluorinated analog of dipalmitoylphosphatidylcholine (DPPC). Surprisingly, we found that the phase transition temperature (Tm) of F-DPPC occurs near 50 degrees C, approximately 10 degrees C higher than its nonfluorinated counterpart, DPPC, as judged by both differential scanning calorimetry and infrared spectroscopy. The pretransition observed for DPPC is absent in F-DPPC. A combination of REDOR, rotational-echo double-resonance, and conventional solid-state NMR experiments demonstrates that F-DPPC forms a fully interdigitated bilayer in the gel phase. Electron paramagnetic resonance experiments show that below Tm, the hydrocarbon chains of F-DPPC are more motionally restricted than those of DPPC. X-ray scattering experiments confirm that the thickness and packing of gel phase F-DPPC is similar to that of heptanetriol-induced interdigitated DPPC. F-DPPC is the first phosphoglyceride containing sn-1 and sn-2 ester-linked fatty acyl chains of equal length that spontaneously forms interdigitated bilayers in the gel state in the absence of inducing agents such as alcohols.  相似文献   
946.
1. Alternative farming practices such as set-aside and agroforestry are likely to be of continuing interest to European agriculture but may have associated problems, such as increased populations of crop pests such as slugs.
2. A silvoarable agroforestry experiment has been in progress since 1987 at Leeds University Farms at Bramham, West Yorkshire, UK. It consists of four replicate blocks, each with rows of trees separating alleys of arable crops; all four blocks have their own arable control areas in adjacent fields.
3. Pitfall trap catches within the experiment indicate that the slug population increased over the period 1991–94. The increase was greatest, and most consistent, within the tree rows in the agroforestry blocks. The increase was slower and less consistent in the arable controls and the arable areas within the agroforestry blocks.
4. In spring 1994, the slugs in each of the treatments in the agroforestry experimental area were sampled using pipe traps, refuge traps and pitfall traps. The number and diversity of slugs were highest in the grassed understorey beneath the rows of trees and significantly higher in the alleys between the rows of trees than in the arable control areas.
5. The levels of slug damage to a pea crop were assessed by surveys that recorded the number of emerging plants and the number of damaged leaves per plant. There were significant correlations between the number of slugs caught and the damage to the crop by slugs. It is concluded that slugs have the potential to be important pests of some crops in silvoarable agroforestry landscapes and that this could influence the choice of crops for this type of farming.
6. Major conclusions are emboldened in the Discussion.  相似文献   
947.
1. This study addresses the issue of structure in sperm whale ( Physeter macrocephalus Linnaeus) populations and whether it is geographically based.
2. During a survey around the South Pacific Ocean, we collected sloughed skin for genetic analyses, recorded coda vocalizations, and photographed fluke markings.
3. Groups of female and immature sperm whales had characteristic mitochondrial haplotypes, coda repertoires, and fluke-mark patterns, but there was no clear geographical structure in any of these attributes.
4. However, similarities of coda repertoire and mitochondrial haplotype distribution were significantly correlated among pairs of groups in a manner that was not geographically based. There was also a significant canonical correlation coefficient between coda repertoire and fluke-mark patterns.
5. These results suggest that attributes (such as vocal repertoire and techniques of predator defence) which are acquired matrilineally, and probably culturally, are conserved during the fission and dispersal of groups.  相似文献   
948.
We have exploited a cross-species expression screen to search for cellular immortalizing activities. A newt blastemal cDNA expression library was transfected into rat embryo fibroblasts and immortal cell lines were selected. This identified a 1-kb cDNA fragment which has a low representation in the cDNA library and is derived from the 3′-UTR of an α-glucosidase-related mRNA. Expression of this sequence in rat embryo fibroblasts has shown that it is active in promoting colony formation and immortalization. It is also able to cooperate with an immortalization-defective deletion mutant of SV40 T antigen, indicating that it can exert its growth-stimulatory activity in the pathway activated by a viral immortalizing oncogene. This is the first example of an immortalizing activity mediated by an RNA sequence, and further analysis of its mechanism should provide new insights into senescence and immortalization.  相似文献   
949.
Small-subunit ribosomal DNA (SSU rDNA) from 20 phenotypically distinct strains of 2,4-dichlorophenoxyacetic acid (2,4-D)-degrading bacteria was partially sequenced, yielding 18 unique strains belonging to members of the alpha, beta, and gamma subgroups of the class Proteobacteria. To understand the origin of 2,4-D degradation in this diverse collection, the first gene in the 2,4-D pathway, tfdA, was sequenced. The sequences fell into three unique classes found in various members of the beta and gamma subgroups of Proteobacteria. None of the α-Proteobacteria yielded tfdA PCR products. A comparison of the dendrogram of the tfdA genes with that of the SSU rDNA genes demonstrated incongruency in phylogenies, and hence 2,4-D degradation must have originated from gene transfer between species. Only those strains with tfdA sequences highly similar to the tfdA sequence of strain JMP134 (tfdA class I) transferred all the 2,4-D genes and conferred the 2,4-D degradation phenotype to a Burkholderia cepacia recipient.Bacteria capable of mineralizing 2,4-dichlorophenoxyacetic acid (2,4-D), a commonly used herbicide, are found in many different phylogenetic groups (2, 3, 7, 11, 22, 23). Evidence suggests that numerous variants of 2,4-D catabolic genes exist and that catabolic operons consist of a near-random mixing of these variants (7). Interspecies gene transfer is a well-documented phenomenon (13), and horizontal gene transfer of the 2,4-D-degrading plasmid pJP4 has been shown (3, 5). However, not all 2,4-D catabolic operons are found on plasmids (10, 11, 16, 20). The extent to which other 2,4-D genes have been exchanged in nature is unknown. The aim of this research was to assess the role of horizontal gene transfer in the evolution of 2,4-D-degrading strains. This article summarizes the results of two aspects of this work—the study of the transfer of the entire 2,4-D pathway by using standard mating experiments and a phylogenetic study of the tfdA gene. The tfdA gene codes for an α-ketoglutarate-dependent 2,4-D dioxygenase which converts 2,4-D into 2,4-dichlorophenol and glyoxylate (6). This 861-bp gene was first sequenced from Ralstonia eutropha JMP134 (19). Two more tfdA genes were cloned from chromosomal locations in Burkholderia strain RASC and Burkholderia strain TFD6 (16, 20). These proved to be identical to each other and 78.5% similar to the original. An alignment of the two variants allowed conserved areas to be identified and primers to be designed for the amplification of tfdA-like genes from other sources (24). Sequence analysis of putative tfdA fragments and the small-subunit ribosomal DNA (SSU rDNA) of the strains carrying them allowed us to construct phylogenies of the genes and their hosts and to look for congruency between them.

Mating experiments.

A collection of 2,4-D degraders containing 15 unique strains as determined by genomic fingerprinting (7) was used as a source of donors in a series of mating experiments (Table (Table1).1). Burkholderia cepacia D5, lacking the ability to grow on 2,4-D and not hybridizing to any tfd genes, was used as a recipient in mating experiments. Strain D5 contains neomycin phosphotransferase genes (nptII) carried on transposon Tn5 and is resistant to 50 μg each of kanamycin, carbenicillin, and bacitracin per ml. All of the 2,4-D strains used were sensitive to these antibiotics. Filter matings were performed with a donor-to-recipient ratio of 1:10. Colonies which grew on selective medium (500 ppm of 2,4-D in mineral salts agar [MMO] [23] including 50 μg of kanamycin, carbenicillin, and bacitracin per ml) were subjected to further tests. Their ability to catabolize 2,4-D was tested in liquid medium (same composition as that described above).

TABLE 1

2,4-D-degrading strains, geographic origins, and GenBank accession numbers
StrainGenBank accession no. (SSU rDNA)OriginMost similar to genus and/or speciesaTransferbtfdA typecGenBank accession no. (tfdA gene)Reference or source
JMP134AF049542AustraliaRalstonia eutropha+IM167303
EML1549AF049546OregonBurkholderia sp.+I2
TFD39AF049539SaskatchewanBurkholderia sp.+IU4319723
K712AF049543MichiganBurkholderia sp.+IU4327611
TFD9AF049537SaskatchewanAlcaligenes xylosoxidans+IU4327623
TFD41AF049541MichiganRalstonia eutropha+I23
TFD38AF049540MichiganRalstonia eutropha+NDc23
TFD23AF049536MichiganRhodoferax fermentans+IU4327623
RASCAF049544OregonBurkholderia sp.(+)IIU257172
TFD6AF049546MichiganBurkholderia sp.II23
TFD2AF049545MichiganBurkholderia sp.II23
TFD31AF049536SaskatchewanRhodoferax fermentansIII23
B6-9AF049538OntarioRhodoferax fermentansNDIIIU431969
I-18U22836OregonHalomonas sp.NDIIIU2249915
K1443AF049531MichiganSphingomonas sp.d11
2,4-D1AF049535MontanaSphingomonas sp.R. Sanford
B6-5AF049533OntarioSphingomonas sp.ND9
B6-10AF049534OntarioSphingomonas sp.ND9
EML146AF049532OregonSphingomonas sp.2
M1AF049530French PolynesiaRhodospeudomonas sp.NDR. Fulthorpe
Open in a separate windowaThe generus and/or species most similar to the strain is given based on similarities of SSU rDNA sequences. bSymbols: +, able to transfer 2,4-D degradation to B. cepacia D5; (+), able to transfer at very low frequency; −, no transfer detected. cND, not determined. d—, no amplificate was obtained. The disappearance of 2,4-D from the culture medium was monitored by high-performance liquid chromatography. Cells were removed by centrifugation, and the supernatant was filtered through 0.2-μm-pore-size filters. These samples were then analyzed on a Lichrosorb Rp-18 column (Anspec Co., Ann Arbor, Mich.) with 60% methanol–40% 0.1% H3PO4 as the eluant. 2,4-D was detected by measuring light absorption at 230 nm. The presence of tfd genes was detected by hybridizing colony blots with a DNA probe derived from the entire pJP4 plasmid. The identity of the colonies was confirmed by probing with the nptII gene of Tn5 (found in B. cepacia D5). Probes were labeled with random hexanucleotides incorporating [32P]dCTP (3,000 Ci/mmol; New England Nuclear, Boston, Mass.). Hybridizations were done under high-stringency conditions by using 50% formamide and Denhardt’s solution (18) at 42°C. Of the 15 unique strains tested, 9 transferred 2,4-D degradation abilities to D5. This transfer was confirmed by hybridization with pJP4 for eight of these strains. B. cepacia RASC could transfer degradative abilities, but neither it nor the transconjugant hybridized to the pJP4 probe. Work subsequent to this study has confirmed that the genes carried by RASC do not hybridize to those found on pJP4 under high-stringency conditions (7).

Phylogenetic analyses.

Total genomic DNA was isolated from 20 unique 2,4-D-degrading strains (including all 15 used for mating experiments) grown on 500 ppm of 2,4-D mineral salts medium amended with 50 ppm of yeast extract. SSU rDNA was amplified by using fD1 and rD1 as primers (25). Putative tfdA fragments were amplified by using primers TVU and TVL as previously described (24). PCR products were purified with a Gene Clean kit (Bio 101, La Jolla, Calif.). Sequencing was done with an Applied Biosystems model 373A automatic sequencer (Perkin-Elmer Cetus) by using fluorescently labeled dye termination at the Michigan State University Sequencing Facility. The sequencing primer used for SSU rDNA fragments was 519R (5′ GTA TTA CCG CGG CTG CTG G-3′). For tfdA fragments, the sequencing primers were the same as the amplification primers. GenBank accession numbers for these sequences are given in Table Table11.The SSU rDNA sequences were compared to sequences in GenBank by using the Basic Local Alignment Search Tool (BLAST) (1), and those strains with the highest maximal segment pair scores were retrieved from GenBank and included in the phylogenetic analysis. Sequences were aligned manually with the software SeqEd (Applied Biosystems) and with MacClade (14). Sites where nucleotides were not resolved for all sequences were deleted from the alignment, as were those nucleotides corresponding to the small loop in this region that is absent in the alpha subgroup of the class Proteobacteria. These deletions left 283 unambiguous sites for the construction of the SSU rDNA phylogenies. Phylogenetic trees were constructed by using the neighbor-joining analysis of pairwise Jukes-Cantor distances (4), and the topology was confirmed by using the maximum parsimony method PAUP (21). Desulfomonile tiedjei of the δ-Proteobacteria was used as an outgroup. Bootstrap analysis based on 100 replicates was used to place confidence estimates on the tree. Only bootstrap values of greater than 50 were used.

2,4-D degrader diversity.

The 2,4-D degraders in this study were distributed throughout the alpha, beta, and gamma subgroups of the Proteobacteria (Fig. (Fig.1).1). The lack of representation of gram-positive bacteria is likely a reflection of isolation methods, not of the lack of gram-positive 2,4-D degraders. The majority of these strains were members of the beta subgroup of Proteobacteria, five of which were most closely related to the genus Burkholderia, having at least 92% sequence similarity with each other. Three were closely related to Rhodoferax fermentans (close to the class Comamonadaceae), three were related to Ralstonia eutropha, and one was related to Alcaligenes xylosoxidans. TFD39 falls outside any clear cluster. One member of the γ-Proteobacteria, strain I-18, a haloalkaliphile, was found to be closely related to the salt-loving genus Halomonas (15). The remaining six strains all clustered in the alpha branch of Proteobacteria (Fig. (Fig.1).1). Of this subgroup, five were most closely related to the genus Sphingomonas. One member of the α-Proteobacteria, strain M1, which is the most oligotrophic and slow growing of all the strains used in this study, is 97% similar to Rhodopseudomonas palustris. The character of strain M1 correlates well with its phylogenetic placement near the slow-growing genus Bradyrhizobium. Open in a separate windowFIG. 1Neighbor-joining dendrogram (Jukes-Cantor distances) of SSU rDNA from 2,4-D-degrading bacteria (indicated in boldface type) and reference strains (indicated in italic type). Class I (•), class II (▴), and class III (■) types of tfdA genes are indicated. Bootstrap confidence limits (percentages) are indicated above each branch. Scale bar represents a Jukes-Cantor distance of 0.01.

tfdA gene fragments.

tfdA gene fragments were successfully amplified and sequenced from 10 strains of β-Proteobacteria and 1 strain of γ-Protobacteria. None of the strains from the α-Proteobacteria gave any amplificates with these primers. These 313 contiguous nucleotides were aligned with additional tfdA sequences from JMP134 and from strain RASC (Fig. (Fig.2).2). Three distinct classes of tfdA gene sequences with slight variations in each class were found. Class I included fragments from JMP134, TFD39, TFD23, K712, and TFD9 that differed from each other by 2 bp at the most. Class I tfdA genes are probably plasmid encoded. All strains with a class I tfdA gene examined so far contained broad-host-range, self-transmissible plasmids containing 2,4-D genes (2, 3, 11, 17). All of the strains with a class I tfdA gene were able to transfer the 2,4-D phenotype in the mating studies reported above. The class II tfdA sequences included identical fragments amplified from RASC, TFD6, and TFD2 which were 76% similar to those in class I. Class III included identical fragments from strains TFD31, B6-9, and I-18 which were 77% similar to class I genes and 80% similar to class II genes. Both class II and III tfdA genes differed from each other and from class I genes in the same nine sites corresponding to the third base pair of the codons. The tfdA phylogenetic tree is a simple one, with three distinct branches that are incongruent with the SSU rDNA-derived phylogeny (Fig. (Fig.3).3). Class I tfdA sequences were found in Burkholderia-like strains, in strains related to the Comamonas-Rhodoferax group, and in the Ralstonia-Acaligenes group, all in the β-Proteobacteria. Class II sequences are less widely distributed, found only in Burkholderia-like branches. However, even in this subgroup, this tfdA variant is found in strains that differ by 7% at the SSU rDNA level (RASC and TFD2). However, the class III sequences were most interesting, being found both in the Comamonas-Rhodoferax group and in a strain of the γ-Proteobacteria, I-18, strains that differ by 24% at the SSU rDNA level. Class III genes have since been found in a collection of randomly isolated non-2,4-D degraders, including gram-positive bacilli, as well as in various gram-negative bacteria, even though the gene is not expressed (10). Open in a separate windowFIG. 2Alignment of 313 nucleotides of internal fragments of tfdA genes from representative strains. Nucleotides identical to tfdA from pJP4 are represented by periods.Open in a separate windowFIG. 3Phylogenetic incongruency of tfdA genes and SSU rDNA from diverse 2,4-D-degrading bacteria. Dendrograms for tfdA and SSU rDNA are indicated. Shading indicates the type of tfdA sequence, either class I, II, or III. Note that branch lengths are not drawn to scale.An interesting result was the detection of two different tfdA gene variants in sibling strains. TFD23 and TFD31 are identical at the ribosomal gene level, but one harbors a class I gene and the other harbors a class III gene. Similarly, TFD6 and EML159 are rRNA siblings that carry a class II and class I gene, respectively.None of the α-Proteobacteria yielded a PCR product when amplified with the conserved tfdA primers. This finding complements our observation that none of these bacteria hybridized to the tfdA gene, even under conditions of low stringency, indicating that any tfdA-like genes in the α-Proteobacteria are likely to be more divergent from the ones sequenced here (7, 11). In addition, none of the Sphingomonas strains in the study hybridized with a whole pJP4 probe, and similarly, no Sphingomonas strains scored positive for transfer of 2,4-D-degrading ability to recipient B. cepacia D5. Together these results suggest a reduced gene flow between members of the α- and β- or γ-Proteobacteria or poor gene expression of β- or γ-derived genes by α-Proteobacteria. Although plasmid pJP4 is a broad-host-range plasmid and has been known to transfer to α-Proteobacteria such as Rhizobium and Agrobacterium species and to γ-Proteobacteria such as Pseudomonas putida, Pseudomonas fluorescens, and Pseudomonas aeruginosa, the 2,4-D pathway is not expressed in these strains of the α- or γ-Proteobacteria (3). Phylogenetically limited expression of plasmid-borne 3-chlorobenzoate-degradative genes has also been noted for the pseudomonads (8). Subsequent studies have found divergent but related sequences for the tfdB and tfdC genes in 2,4-D-degrading Sphingomonas strains (7, 12, 24).With the exceptions of the minor differences within the class I pJP4-like tfdA sequences, there were no intermediate tfdA sequences. The most likely explanation of this is that the rate of horizontal transfer of the tfd genes is high relative to the rate at which mutations can accumulate. Examination of sequences of tfdA genes from a greater variety of organisms may turn up more intermediate variation.  相似文献   
950.
The influence of vesicular–arbuscular mycorrhizal (M) colonization on biomass production and photosynthesis of Trifolium repens L. was investigated in two experiments in which the foliar nitrogen and phosphorus contents of non-mycorrhizal (NM) plants were manipulated to be no lower than that of M plants. Throughout both experiments there was a stimulation in the rate of CO2 assimilation of the youngest, fully expanded leaf of M compared with NM plants. In addition, M plants exhibited a higher specific leaf area compared with NM plants, a response that maximized the area available for CO2 assimilation per unit of carbon (C) invested. Despite the increased rate of photosynthesis in M plants there was no evidence that the additional C gained was converted to biomass production of M plants. It is suggested that this additional C gained by colonized plants was allocated to the mycorrhizal fungus and that it is the fungus, by acting as a sink for assimilates, that facilitated the stimulation in the rate of photosynthesis of the plant partner.  相似文献   
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