全文获取类型
收费全文 | 1614篇 |
免费 | 187篇 |
出版年
2022年 | 9篇 |
2021年 | 30篇 |
2020年 | 14篇 |
2019年 | 14篇 |
2018年 | 13篇 |
2017年 | 11篇 |
2016年 | 23篇 |
2015年 | 65篇 |
2014年 | 53篇 |
2013年 | 84篇 |
2012年 | 87篇 |
2011年 | 92篇 |
2010年 | 45篇 |
2009年 | 47篇 |
2008年 | 90篇 |
2007年 | 77篇 |
2006年 | 85篇 |
2005年 | 72篇 |
2004年 | 58篇 |
2003年 | 76篇 |
2002年 | 59篇 |
2001年 | 60篇 |
2000年 | 62篇 |
1999年 | 54篇 |
1998年 | 24篇 |
1997年 | 21篇 |
1996年 | 19篇 |
1995年 | 23篇 |
1994年 | 25篇 |
1993年 | 21篇 |
1992年 | 27篇 |
1991年 | 22篇 |
1990年 | 35篇 |
1989年 | 28篇 |
1988年 | 28篇 |
1987年 | 18篇 |
1986年 | 15篇 |
1985年 | 18篇 |
1984年 | 17篇 |
1983年 | 16篇 |
1982年 | 9篇 |
1981年 | 13篇 |
1979年 | 20篇 |
1978年 | 15篇 |
1977年 | 13篇 |
1976年 | 10篇 |
1975年 | 8篇 |
1974年 | 8篇 |
1973年 | 12篇 |
1972年 | 8篇 |
排序方式: 共有1801条查询结果,搜索用时 31 毫秒
961.
962.
963.
Blood samples from 94 coal tits (Parus ater), 56 great tits (Parus major) and 219 pied flycatchers (Ficedula hypoleuca), caught between 1993 and 2002 at two localities in Lower Saxony, Germany, were examined for haemosporidian infection by
parasite-specific polymerase chain reaction (PCR). A simple PCR targeting the 18 SSU rRNA gene of the parasites was used for
rapid screening of the samples and generated a total infection prevalence of 20.6% (76/369): 6.8% (n = 15) of the pied flycatchers, 19.1% (n = 18) of the coal tits and 76.8% (n = 43) of the great tits were infected. The positive specimens were re-examined by a cytochrome b gene-directed nested PCR producing significantly longer DNA fragments (approx. 520 bp) that were sequenced and analysed against
GenBank-deposited nucleotide sequences. In various numbers (once to 30 times), a total of 13 parasitic DNA sequences differing
from 2.9 to 8.5% (13–45 nucleotides) were demonstrated in the three bird species. Due to similarities of 98–100% with GenBank
entries, 11 sequences could be assigned to Plasmodium sp. and two to the genus Haemoproteus. In summary, 57 birds were infected with Plasmodium and 19 with Haemoproteus, corresponding to 15.4 and 5.1% of all birds examined, and to 75 and 25% of all birds tested positive. As the only defined
species, Haemoproteus majoris was identified in 17 great tits. 相似文献
964.
Winther-Larsen HC Wolfgang MC van Putten JP Roos N Aas FE Egge-Jacobsen WM Maier B Koomey M 《Journal of bacteriology》2007,189(18):6676-6685
Type IV pili (TFP) play central roles in the expression of many phenotypes including motility, multicellular behavior, sensitivity to bacteriophages, natural genetic transformation, and adherence. In Neisseria gonorrhoeae, these properties require ancillary proteins that act in conjunction with TFP expression and influence organelle dynamics. Here, the intrinsic contributions of the pilin protein itself to TFP dynamics and associated phenotypes were examined by expressing the Pseudomonas aeruginosa PilA(PAK) pilin subunit in N. gonorrhoeae. We show here that, although PilA(PAK) pilin can be readily assembled into TFP in this background, steady-state levels of purifiable fibers are dramatically reduced relative those of endogenous pili. This defect is due to aberrant TFP dynamics as it is suppressed in the absence of the PilT pilus retraction ATPase. Functionally, PilA(PAK) pilin complements gonococcal adherence for human epithelial cells but only in a pilT background, and this property remains dependent on the coexpression of both the PilC adhesin and the PilV pilin-like protein. Since P. aeruginosa pilin only moderately supports neisserial sequence-specific transformation despite its assembly proficiency, these results together suggest that PilA(PAK) pilin functions suboptimally in this environment. This appears to be due to diminished compatibility with resident proteins essential for TFP function and dynamics. Despite this, PilA(PAK) pili support retractile force generation in this background equivalent to that reported for endogenous pili. Furthermore, PilA(PAK) pili are both necessary and sufficient for bacteriophage PO4 binding, although the strain remains phage resistant. Together, these findings have significant implications for TFP biology in both N. gonorrhoeae and P. aeruginosa. 相似文献
965.
Peptide transport in Helicobacter pylori: roles of dpp and opp systems and evidence for additional peptide transporters 下载免费PDF全文
Despite research into the nutritional requirements of Helicobacter pylori, little is known regarding its use of complex substrates, such as peptides. Analysis of genome sequences revealed putative ABC-type transporter genes for dipeptide (dppABCDF) and oligopeptide (oppABCD) transport. Genes from each system were PCR amplified, cloned, and disrupted by cassette insertion either individually (dppA, dppB, dppC, oppA, oppB, and oppC) or to create double mutants (dppA oppA, dppB oppB, dppB dppC, and oppB oppC). Peptide-utilizing abilities of the strains were assessed by monitoring growth in a chemically defined medium where the only source of the essential amino acid isoleucine was from peptides of various lengths (two to nine amino acids long). The dipeptide system mutants lacked the ability to use certain dipeptides, hexapeptides, and nonapeptides. However, these mutants retained some ability to grow with other dipeptides, tripeptides, and tetrapeptides. Of the oligopeptide mutants, only the oppB strain differed significantly from the wild type. This strain showed a wild-type phenotype for growth with longer peptides (hexa- and nonapeptides) while having a decreased ability to utilize di-, tri-, and tetrapeptides. The dppA oppA and dppB oppB mutants showed similar phenotypes to those of the dppA and dppB mutants, respectively. Peptide digestion by metalloproteases was ruled out as the cause for residual peptide transport by growing mutant strains in the presence of either EDTA or EGTA. Degradation products associated with a fluorescein isothiocyanate-labeled hexapeptide (plus cells) were minimal. An as yet unidentified peptide transport system(s) in H. pylori is proposed to be responsible for the residual transport. 相似文献
966.
In ionotropic glutamate receptors, agonist binding occurs in a conserved clam shell-like domain composed of the two lobes D1 and D2. Docking of glutamate into the binding cleft promotes rotation in the hinge region of the two lobes, resulting in closure of the binding pocket, which is thought to represent a prerequisite for channel gating. Here, we disrupted D1D2 interlobe interactions in the NR2A subunit of N-methyl-d-aspartate (NMDA) receptors through systematic mutation of individual residues and studied the influence on the activation kinetics of currents from NR1/NR2 NMDA receptors heterologously expressed in HEK cells. We show that the mutations affect differentially glutamate binding and channel gating, depending on their location within the binding domain, mainly by altering k(off) and k(cl), respectively. Whereas impaired stability of glutamate in its binding site is the only effect of mutations on one side of the ligand binding pocket, close to the hinge region, alterations in gating are the predominant consequence of mutations on the opposite side, at the entrance of the binding pocket. A mutation increasing D1D2 interaction at the entrance of the pocket resulted in an NMDA receptor with an increased open probability as demonstrated by single channel and whole cell kinetic analysis. Thus, the results indicate that agonist-induced binding domain closure is itself a complex process, certain aspects of which are coupled either to binding or to gating. Specifically, we propose that late steps of domain closure, in kinetic terms, represent part of channel gating. 相似文献
967.
Melkonian KA Maier KC Godfrey JE Rodgers M Schroer TA 《The Journal of biological chemistry》2007,282(27):19355-19364
Dynamitin is a commonly used inhibitor of cytoplasmic dynein-based motility in living cells. Dynamitin does not inhibit dynein directly but instead acts by causing disassembly of dynactin, a multiprotein complex required for dynein-based movement. In dynactin, dynamitin is closely associated with the subunits p150(Glued) and p24, which together form the shoulder and projecting arm structures of the dynactin molecule. In this study, we explore the way in which exogenous dynamitin effects dynactin disruption. We find that pure, recombinant dynamitin is an elongated protein with a strong propensity for self-assembly. Titration experiments reveal that free dynamitin binds dynactin before it causes release of subunits. When dynamitin is added to dynactin at an equimolar ratio of exogenous dynamitin subunits to endogenous dynamitin subunits (1x= 4 mol of exogenous dynamitin per mole of dynactin), exogenous dynamitin exchanges with endogenous dynamitin, and partial release of p150(Glued) is observed. When added in vast excess (> or =25x; 100 mol of exogenous dynamitin per mole of dynactin), recombinant dynamitin causes complete release of both p150(Glued) subunits, two dynamitins and one p24, but not other dynactin subunits. Our data suggest that dynamitin mediates disruption of dynactin by binding to endogenous dynamitin subunits. This binding destabilizes the shoulder structure that links the p150(Glued) arm to the Arp1 filament and leads to subunit release. 相似文献
968.
Maier CT 《Journal of economic entomology》2007,100(4):1291-1297
The distribution and hosts of the exotic cedar-boring beetle, Callidiellum rufipenne (Motschulsky) (Coleoptera: Cerambycidae), were determined in five northeastern U.S. states by capturing adults on cedar trap logs and by rearing adults from various conifers. This beetle was detected in the coastal states of Massachusetts, Rhode Island, Connecticut, New York, and New Jersey. In these states, adults emerged from the live or dead wood of four genera and eight species of Cupressaceae; species of Pinaceae were not hosts. Through its entire range, C. rufipenne is reported to infest at least 14 species of Cupressaceae, four species of Pinaceae, and one species of Taxaceae; but, records of Pinaceae and possibly Taxaceae are suspect. Based on the number of adults that emerged from coniferous poles in a five-way choice test in the field, the infestation level was significantly greater in Chamaecyparis thyoides (L.) Britton, Sterns, and Poggenburg and Juniperus virginiana L. than in Pinus rigida Miller, Pinus strobus L., and Tsuga canadensis (L.) Carribre (last three species uninfested). In a second test of host preference in the wild, beetles infested four cupressaceous species, but not Abies balsamea (L.) Miller, Picea rubens Sargent, Pinus rigida, P. strobus, and Ts. canadensis in the Pinaceae. Infestation level was highest in Ch. thyoides, followed in decreasing order by Juniperus communis L., Thuja occidentalis L., and J. virginiana. In a comparison of live and dead J. virginiana, beetles developed to adults only in dead trees (36 beetles per tree). When trunk sections of Th. occidentalis with and without bark were offered to females in cages, beetles of the next generation emerged exclusively from wood with bark. In the Northeast, only species of Cupressaceae apparently are suitable hosts for C. rufipenne. Infestation of these species may be prevented or reduced by proper care of live plants and by debarking trees after harvesting. 相似文献
969.
MutY is an adenine glycosylase that has the ability to efficiently remove adenines from adenine/7,8-dihydro-8-oxoguanine (8-oxo-G) or adenine/guanine mismatches, and plays an important role in oxidative DNA damage repair. The human gastric pathogen Helicobacter pylori has a homolog of the MutY enzyme. To investigate the physiological roles of MutY in H. pylori, we constructed and characterized a mutY mutant. H. pylori mutY mutants incubated at 5% O2 have a 325-fold higher spontaneous mutation rate than its parent. The mutation rate is further increased by exposing the mutant to atmospheric levels of oxygen, an effect that is not seen in an E. coli mutY mutant. Most of the mutations that occurred in H. pylori mutY mutants, as examined by rpoB sequence changes that confer rifampicin resistance, are GC to TA transversions. The H. pylori enzyme has the ability to complement an E. coli mutY mutant, restoring its mutation frequency to the wild-type level. Pure H. pylori MutY has the ability to remove adenines from A/8-oxo-G mismatches, but strikingly no ability to cleave A/G mismatches. This is surprising because E. coli MutY can more rapidly turnover A/G than A/8-oxo-G. Thus, H. pylori MutY is an adenine glycosylase involved in the repair of oxidative DNA damage with a specificity for detecting 8-oxo-G. In addition, H. pylori mutY mutants are only 30% as efficient as wild-type in colonizing the stomach of mice, indicating that H. pylori MutY plays a significant role in oxidative DNA damage repair in vivo. 相似文献
970.
Oliver Knapp Elke Maier Eva Waltenberger Christelle Mazuet Roland Benz Michel R. Popoff 《Cellular microbiology》2015,17(2):288-302
Clostridium perfringens iota toxin is a binary toxin that is organized into enzyme (Ia) and binding (Ib) components. Ib forms channels in lipid bilayers and mediates the transport of Ia into the target cells. Here we show that Ib residues 334–359 contain a conserved pattern of alternating hydrophobic and hydrophilic residues forming two amphipathic β‐strands involved in membrane insertion and channel formation. This stretch of amino acids shows remarkable structural and functional analogies with the β‐pore‐forming domain of C. perfringens epsilon toxin. Several mutations within the two amphipathic β‐strands affected pore formation, single‐channel conductance and ion selectivity (S339E‐S341E, Q345H N346E) confirming their involvement in channel formation. F454 of Ib corresponds to the Φ‐clamp F427 of anthrax protective antigen and F428 of C2II binary toxins. The mutation F454A resulted in a loss of cytotoxicity and strong increase in single‐channel conductance (500 pS as compared with 85 pS in 1 M KCl) with a slight decrease in cation selectivity, indicating that the Φ‐clamp is highly conserved and crucial for binary toxin activity. In contrast, the mutants Q367D, N430D, L443E had no or only minor effects on Ib properties, while T360I, T360A and T360W caused a dramatic effect on ion selectivity and single‐channel conductance, indicating gross disturbance of the oligomer structure. This suggests that, at least in the iota toxin family, T360 has a structural role in the pore organization. Moreover, introduction of charged residues within the channel (S339E‐S341E) or in the vestibule (Q367D, N430D and L443E) had virtually no effect on chloroquine or Ia binding, whereas F454A, T360I, T360A and T360W strongly decreased the chloroquine and Ia affinity to Ib. These results support that distinct residues within the vestibule interact with chloroquine and Ia or are responsible for channel structure, while the channel lining amino acids play a less important role. 相似文献