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61.
Inactivation of Human Immunodeficiency Virus Type 1 Infectivity with Preservation of Conformational and Functional Integrity of Virion Surface Proteins 总被引:27,自引:9,他引:18
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62.
Constitutive and tissue-specific differential expression of the cryIA(b) gene in transgenic rice plants conferring resistance to rice insect pest 总被引:11,自引:0,他引:11
K. Datta A. Vasquez J. Tu L. Torrizo M. F. Alam N. Oliva E. Abrigo G. S. Khush S. K. Datta 《TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik》1998,97(1-2):20-30
The truncated chimeric Bt gene, cryIA(b) of Bacillus thuringiensis, driven by two constitutive promoters, 35S from CaMV and Actin-1 from rice, and two tissue-specific promoters, pith tissue
and pepcarboxylase (PEPC) for green tissue from maize, was introduced into several varieties of rice (indica and japonica)
by microprojectile bombardment and protoplast systems. A total of 1800 putative transgenic Bt rice plants could be produced. Southern analysis revealed that more than 100 independently transformed plants could be confirmed
for integration of the cryIA(b) gene. High levels of CryIA(b) proteins were obtained in the green tissue (leaves and stem) of many plants using the PEPC promoter. There was little difference
in Bt protein level in leaves and stems from transgenic plants with the 35 S or Actin-1 promoter. Out of 800 Southern-positive
plants that were bioassayed, 81 transgenic plants showed 100% mortality of insect larvae of the yellow stem borer (Scirpophaga incertulas). The transgene, cryIA(b), driven by different promoters showed a wide range of expression (low to high) of Bt proteins stably inherited in a number
of rice varieties with enhanced yellow stem borer resistance. This first report of transgenic indica Bt rice plants with the PEPC or pith promoter either alone or in combination should provide a better strategy for providing
rice plants with protection against insect pest resistance, minimizing the expression of the CryIA(b) protein in seeds and
other tissues.
Received: 12 November 1997 / Accepted: 25 November 1997 相似文献
63.
Females of the sea urchin Strongylocentrotus purpuratus differ in the structures of their egg jelly sulfated fucans 总被引:1,自引:0,他引:1
The egg jelly coats of sea urchins contain sulfated fucans which bind to a
sperm surface receptor glycoprotein to initiate the signal transduction
events resulting in the sperm acrosome reaction. The acrosome reaction is
an ion channel regulated exocytosis which is an obligatory event for sperm
binding to, and fusion with, the egg. Approximately 90% of individual
females of the sea urchin Strongylocentrotus purpuratus spawned eggs having
only one of two possible sulfated fucan electrophoretic isotypes, a slow
migrating (sulfated fucan I), or a fast migrating (sulfated fucan II)
isotype. The remaining 10% of females spawned eggs having both sulfated
fucan isotypes. The two sulfated fucan isotypes were purified from egg
jelly coats and their structures determined by NMR spectroscopy and
methylation analysis. Both sulfated fucans are linear polysaccharides
composed of 1-->3-linked alpha-L-fucopyranosyl units. Sulfated fucan I
is entirely sulfated at the O -2 position but with a heterogeneous
sulfation pattern at O -4 position. Sulfated fucan II is composed of a
regular repeating sequence of 3 residues, as follows: [3-alpha-L-Fuc p -
2,4(OSO3)-1-->3-alpha-L-Fuc p -4(OSO3)-1-->3-alpha-L-Fuc p -4(OSO3)-
1]n. Both purified sulfated fucans have approximately equal potency in
inducing the sperm acrosome reaction. The significance of two structurally
different sulfated fucans in the egg jelly coat of this species could
relate to the finding that the sperm receptor protein which binds sulfated
fucan contains two carbohydrate recognition modules of the C-type lectin
variety which differ by 50% in their primary structure.
相似文献
64.
Sequence variations in small-subunit ribosomal RNAs of Hartmannella vermiformis and their phylogenetic implications 总被引:1,自引:0,他引:1
Evidence of associations between free-living amoebas and human disease has
been increasing in recent years. Knowledge about phylogenetic relationships
that may be important for the understanding of pathogenicity in the genera
involved is very limited at present. Consequently, we have begun to study
these relationships and report here on the phylogeny of Hartmannella
vermiformis, a free-living amoeba that can harbor the etiologic agent of
Legionnaires' disease. Our analysis is based on studies of small-subunit
ribosomal RNA genes (srDNA). Nucleotide sequences were determined for
nuclear srDNA from three strains of H. vermiformis isolated from the United
Kingdom, Germany, and the United States. These sequences then were compared
with a sequence previously obtained for a North American isolate by J. H.
Gunderson and M. L. Sogin. The four genes are 1,840 bp long, with an
average GC content of 49.6%. Sequence differences among the strains range
are 0.38%-0.76%. Variation occurs at 19 positions and includes 2
single-base indels plus 14 monotypic and 3 ditypic single-base
substitutions. Variation is limited to eight helix/loop structures
according to a current model for srRNA secondary structure. Parsimony,
distance, and bootstrap analyses used to examine phylogenetic relationships
between the srDNA sequences of H. vermiformis and other eukaryotes
indicated that Hartmannella sequences were most closely related to those of
Acanthamoeba and the alga Cryptomonas. All ditypic sites were consistent
with a separation between European and North American strains of
Hartmannella, but results of other tests of this relationship were
statistically inconclusive.
相似文献
65.
Morphine in doses of 1, 2, and 4 mg/kg i.v. produced dose related elevations in cat body temperature while doses of 0.25 and 0.50 mg/kg had no such effect. Tolerance was found to develop to the hyperthermic response after seven days of daily morphine injection. Pretreatment with naloxone at a dose one-fourth the dose of morphine prevented the morphine induced rise in body temperature in all cats tested. When the cats received naloxone after twelve days of daily morphine a withdrawal syndrome resulted and was accompanied by a hypothermia that was proportional to the morphine maintenance dose and severity of withdrawal. 相似文献
66.
Barbara Vasquez Fukashi Ishibashi Barbara V. Howard 《In vitro cellular & developmental biology. Plant》1982,18(7):643-649
Summary Methods have been developed for measuring the intracellular water space (ICS) in cultured diploid fibroblast monolayers. The
values obtained have been compared to similar measurements of ICS in suspended fibroblasts using an oil spin method. Markers
commonly used for measurement of total water space (TWS) ([3H]H2O, [14C]urea) and extracellular space (ECS) ([3H]inulin, [14C]l-glucose, [3H]sucrose, [3H]d-mannitol) were investigated for use in cell monolayers. Monolayer incubations were terminated by rapidly rinsing the culture
dishes three times with ice cold buffer. The distribution volume of the TWS marker [14C]urea plateaued at 10 to 15 min and was independent of urea concentration. [3H]H2O was not a suitable marker for measuring total water space in cell monolayers because of rapid loss of intracellular label
during rinsing. Intracellular space was calculated by subtracting [3H]sucrose space (5 min) from [14C]urea space (20 min) after incubation of fibroblasts with both markers. Values obtained for ICS (mean ± SE;μl/106 cells) of fibroblasts from two cell lines measured in monolayer (1.74±0.11 and 1.60±0.10) and in suspension (1.88±0.07 and
1.78±0.11) was approximately 10% smaller than the values for cell size (2.01 and 2.22) obtained from Coulter Counter sizing.
Thus, the methods developed for measurement of ICS in monolayer fibroblasts yield data comparable to those obtained with the
more standard oil spin method. Furthermore, the methods can be applied to measurement of ICS in other types of adherent cells.
This work was supported in part by funds from the National Institutes of Health Contract N01-AM-9212. 相似文献
67.
Zhao‐chang Fan Lin Shan Benjamin Z. Goldsteen Luke W. Guddat Archana Thakur Nicholas F. Landolfi Man Sung Co Maximiliano Vasquez Cary Queen Paul A. Ramsland Allen B. Edmundson 《Journal of molecular recognition : JMR》1999,12(1):19-32
The objective of this work is to compare the three‐dimensional structures of “humanized” and mouse–human chimeric forms of a murine monoclonal antibody elicited against human γ‐interferon. It is also to provide structural explanations for the small differences in the affinities and biological interactions of the two molecules for this antigen. Antigen‐binding fragments (Fabs) were produced by papain hydrolysis of the antibodies and crystallized with polyethylene glycol (PEG) 8000 by nearly identical microseeding procedures. Their structures were solved by X‐ray analyses at 2.9 Å resolution, using molecular replacement methods and crystallographic refinement. Comparison of these structures revealed marked similarities in the light (L) chains and near identities of the constant (C) domains of the heavy (H) chains. However, the variable (V) domains of the heavy chains exhibited substantial differences in the conformations of all three complementarity‐determining regions (CDRs), and in their first framework segments (FR1). In FR1 of the humanized VH, the substitution of serine for proline in position 7 allowed the N‐terminal segment (designated strand 4‐1) to be closely juxtaposed to an adjacent strand (4‐2) and form hydrogen bonds typical of an antiparallel β‐pleated sheet. The tightening of the humanized structure was relayed in such a way as to decrease the space available for the last portion of HFR1 and the first part of HCDR1. This compression led to the formation of an α‐helix involving residues 25–32. With fewer steric constraints, the corresponding segment in the chimeric Fab lengthened by at least 1 Å to a random coil which terminated in a single turn of 310 helix. In the humanized Fab, HCDR1, which is sandwiched between HCDR2 and HCDR3, significantly influenced the structures of both regions. HCDR2 was forced into a bent and twisted orientation different from that in the chimeric Fab, both at the crown of the loop (around proline H52a) and at its base. As in HCDR1, the last few residues of HCDR2 in the humanized Fab were compressed into a space‐saving α‐helix, contrasting with a more extended 310 helix in the chimeric form. HCDR3 in the humanized Fab was also adjusted in shape and topography. The observed similarities in the functional binding activities of the two molecules can be rationalized by limited induced fit adjustments in their structures on antigen binding. While not perfect replicas, the two structures are testimonials to the progress in making high affinity monoclonal antibodies safe for human use. Copyright © 1999 John Wiley & Sons, Ltd. 相似文献
68.
69.
70.
TM?Matthews RK?Duncan M?Zidanic TH?Michael PA?FuchsEmail author 《Journal of comparative physiology. A, Neuroethology, sensory, neural, and behavioral physiology》2005,191(6):491-503
In the inner ear of birds, as in mammals, reptiles and amphibians, acetylcholine released from efferent neurons inhibits hair cells via activation of an apamin-sensitive, calcium-dependent potassium current. The particular potassium channel involved in avian hair cell inhibition is unknown. In this study, we cloned a small-conductance, calcium-sensitive potassium channel (gSK2) from a chicken cochlear library. Using RT-PCR, we demonstrated the presence of gSK2 mRNA in cochlear hair cells. Electrophysiological studies on transfected HEK293 cells showed that gSK2 channels have a conductance of approximately 16 pS and a half-maximal calcium activation concentration of 0.74±0.17 M. The expressed channels were blocked by apamin (IC50=73.3±5.0 pM) and d-tubocurarine (IC50=7.6±1.0 M), but were insensitive to charybdotoxin. These characteristics are consistent with those reported for acetylcholine-induced potassium currents of isolated chicken hair cells, suggesting that gSK2 is involved in efferent inhibition of chicken inner ear. These findings imply that the molecular mechanisms of inhibition are conserved in hair cells of all vertebrates. 相似文献