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1.
Separation, purification, partial characterization and comparison of the heavy and light chains of botulinum neurotoxin types A, B, and E 总被引:15,自引:0,他引:15
Clostridium botulinum produces botulinum neurotoxin (NT) in antigenically distinct forms. When isolated from bacterial cultures type E is a single chain, type B is a mixture of single and two-chain molecules, and type A is essentially a two-chain molecule (Mr approximately 150,000). Protease(s) in the cultures or trypsin nick single-chain NT to the two-chain form. The heavy (Mr approximately 100,000) and light (Mr approximately 50,000) chains of the two-chain molecule remain held together by -S-S-bond(s). The two chains are presumed to have different functions. NT binds to nerve cells via the heavy chain and then light chain enters the cell and blocks release of acetylcholine (Simpson, L. L. (1981) Pharmacol. Rev. 33, 155-188). We nicked single-chain NT to form the two-chain form with trypsin, minimizing secondary cleavages, then separated and purified the heavy and light chains using ion-exchange chromatography. The technique, with minor modifications, is a generalized method for types A, B, and E. These subunit chains (each a single band in sodium dodecyl sulfatepolyacrylamide gel electrophoresis) were analyzed for their complete amino acid compositions. The amino acid contents of the heavy and light chains agreed well with the parent two-chain molecule. This affirms that NT is composed of two chains. The two subunit chains are now usable for amino acid sequence and other studies. Comparison of the amino acid contents indicates more similarity among the light chains than the heavy chains of the three NT types, a similarity that agrees with our published partial amino acid sequences (first 13-18 residues) of these chains. Several (up to 9) different amino acid residues of the heavy chain (which is twice the size of the light chain) are present in double the number of corresponding residues in the light chain. 相似文献
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3.
Krishnan Neeraja M. Raina Sameer Z. Pollock David D. 《Biological procedures online》2004,6(1):180-188
Substitution patterns among nucleotides are often assumed to be constant in phylogenetic analyses. Although variation in the
average rate of substitution among sites is commonly accounted for, variation in the relative rates of specific types of substitution
is not. Here, we review details of methodologies used for detecting and analyzing differences in substitution processes among
predefined groups of sites. We describe how such analyses can be performed using existing phylogenetic tools, and discuss
how new phylogenetic analysis tools we have recently developed can be used to provide more detailed and sensitive analyses,
including study of the evolution of mutation and substitution processes. As an example we consider the mitochondrial genome,
for which two types of transition deaminations (C⇒T and A⇒G) are strongly affected by single-strandedness during replication,
resulting in a strand asymmetric mutation process. Since time spent single-stranded varies along the mitochondrial genome,
their differential mutational response results in very different substitution patterns in different regions of the genome.
Published: September 2, 2004. 相似文献
4.
Inadequate information about the genetic structure of the polyphagous Rhizoctonia solani has made sheath blight resistance breeding a difficult task. To assess the variability in the Indian populations of sheath blight fungus, 18 isolates were collected from different rice growing regions of India and analyzed for virulence and electrophoretic profiles of 13 isozymes. The virulence spectrum of all 18 isolates was examined on susceptible IR50 and tolerant Swarnadhan varieties, based on which the isolates could be grouped as highly virulent, moderately virulent or a virulent. A total of 11 enzyme systems with 153 electrophoretic phenotypes were applied to characterize the genetic variation among the isolates. Cluster analyses based on isozyme patterns resulted in one major cluster comprising 16 virulent isolates, with two a virulent isolates loosely linked to this at 0.13 similarity. Isozyme systems of esterases (both and ) and 6-phosphogluconic dehydrogenase could be used to fingerprint the individual isolates.This revised version was published online in October 2005 with corrections to the Cover Date. 相似文献
5.
Neeraja Vajrala Luis A. Sayavedra-Soto Peter J. Bottomley Daniel J. Arp 《Archives of microbiology》2010,192(11):899-908
Nitrosomonas europaea has a single three-gene operon (nitABC) encoding an iron ABC transporter system (NitABC). Phylogenetic analysis clustered the subunit NitB with Fe3+-ABC transporter permease components from other organisms. The N.
europaea strain deficient in nitB (nitB::kan) grew well in either Fe-replete or Fe-limited media and in Fe-limited medium containing the catecholate-type siderophore,
enterobactin or the citrate-based dihydroxamate-type siderophore, aerobactin. However, the nitB::kan mutant strain was unable to grow in Fe-limited media containing either the hydroxamate-type siderophores, ferrioxamine and
ferrichrome or the mixed-chelating type siderophore, pyoverdine. Exposure of N. europaea cells to a ferrichrome analog coupled to the fluorescent moiety naphthalic diimide (Fhu-NI) led to increase in fluorescence
in the wild type but not in nitB::kan mutant cells. Spheroplasts prepared from N.
europaea wild type exposed to Fhu-NI analog retained the fluorescence, while spheroplasts of the nitB::kan mutant were not fluorescent. NitABC transports intact Fe3+-ferrichrome complex into the cytoplasm and is an atypical ABC type iron transporter for Fe3+ bound to ferrioxamine, ferrichrome or pyoverdine siderophores into the cytoplasm. The mechanisms to transport iron in either
the Fe3+ or Fe2+ forms or Fe3+ associated with enterobactin or aerobactin siderophores into the cell across the cytoplasmic membrane are as yet undetermined. 相似文献
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Namrata Singh Trang T. M. Dang Georgina V. Vergara Dev Mani Pandey Darlene Sanchez C. N. Neeraja Endang M. Septiningsih Merlyn Mendioro Evelyn Mae Tecson-Mendoza Abdelbagi M. Ismail David J. Mackill Sigrid Heuer 《TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik》2010,121(8):1441-1453
8.
CD101 surface expression discriminates potency among murine FoxP3+ regulatory T cells 总被引:2,自引:0,他引:2
Fernandez I Zeiser R Karsunky H Kambham N Beilhack A Soderstrom K Negrin RS Engleman E 《Journal of immunology (Baltimore, Md. : 1950)》2007,179(5):2808-2814
CD4+CD25+FoxP3+ regulatory T cells (Treg) have been shown to be protective in animal models of autoimmunity and acute graft-vs-host disease. However, owing to the functional heterogeneity among CD4+CD25+ T cells, surface markers expressed selectively on functionally active Treg would be useful for purposes of identifying and isolating such cells. We generated a rabbit mAb against murine CD101, a transmembrane glycoprotein involved in T cell activation. Among freshly isolated T cells, CD101 was detected on 25-30% of CD4+CD25+ Treg and approximately 20% of conventional memory T cells. CD101(high) Treg displayed greater in vitro suppression of alloantigen-driven T cell proliferation as compared with CD101(low) Treg. In a model of graft-vs-host disease induced by allogeneic bone marrow transplantation in vivo bioluminescence imaging demonstrated reduced expansion of donor-derived luciferase-labeled conventional T cells in mice treated with CD101(high) Treg, compared with CD101(low) Treg. Moreover, treatment with CD101(high) Treg resulted in improved survival, reduced proinflammatory cytokine levels and reduced end organ damage. Among the CD101(high) Treg all of the in vivo suppressor activity was contained within the CD62L(high) subpopulation. We conclude that CD101 expression distinguishes murine Treg with potent suppressor activity. 相似文献
9.
CH Balachiranjeevi Naik S. Bhaskar V. Abhilash S. Akanksha B. C. Viraktamath M. S. Madhav A. S. Hariprasad G. S. Laha M. S. Prasad S. M. Balachandran C. N. Neeraja M. Satendra Kumar P. Senguttuvel K. B. Kemparaju V. P. Bhadana T. Ram G. Harika H. K. Mahadeva Swamy S. K. Hajira A. Yugander K. Pranathi M. Anila G. Rekha M. B. V. N. Kousik T. Dilip Kumar R. K. Swapnil Archana Giri R. M. Sundaram 《Molecular breeding : new strategies in plant improvement》2015,35(7):1-12
10.
G. Ramkumar K. Srinivasarao K. Madhan Mohan I. Sudarshan A. K. P. Sivaranjani K. Gopalakrishna C. N. Neeraja S. M. Balachandran R. M. Sundaram M. S. Prasad N. Shobha Rani A. M. Rama Prasad B. C. Viraktamath M. S. Madhav 《Molecular breeding : new strategies in plant improvement》2011,27(1):129-135
Rice blast is one of the most devastating diseases affecting the rice crop throughout the world. In molecular breeding for host plant resistance, functional markers are very useful for enhancing the precision and accuracy in marker-assisted selection (MAS) of target gene(s) with minimum effort, time and cost. Pi54 (which was earlier known as Pik h ) is one of the major blast resistance genes and has been observed to show resistance against many isolates of the blast pathogen in India. The gene has been cloned through map-based strategy and encodes a nucleotide-binding site?Cleucine-rich repeat (NBS?CLRR) domain-containing protein. In the present study, we carried out allele mining for this gene and identified a 144-bp insertion/deletion (InDel) polymorphism in the exonic region of the gene. A PCR-based co-dominant molecular marker targeting this InDel, named Pi54 MAS, was developed. Pi54 MAS was observed to perfectly co-segregate with blast resistance in a mapping population with no recombinants. Validation of this marker in 105 genotypes which are either susceptible or resistant to rice blast disease showed that the marker is polymorphic in most of the resistant?Csusceptible genotype combinations and is more accurate than the earlier reported markers for Pi54. Hence this functional, co-dominant marker is suggested for routine deployment in MAS of Pi54 in breeding programs. 相似文献