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91.
Chai YY Rahman RN Illias RM Goh KM 《Journal of industrial microbiology & biotechnology》2012,39(5):731-741
Two genes that encode α-amylases from two Anoxybacillus species were cloned and expressed in Escherichia coli. The genes are 1,518 bp long and encode 506 amino acids. Both sequences are 98% similar but are distinct from other well-known
α-amylases. Both of the recombinant enzymes, ASKA and ADTA, were purified using an α-CD–Sepharose column. They exhibited an
optimum activity at 60°C and pH 8. Both amylases were stable at pH 6–10. At 60°C in the absence of Ca2+, negligible reduction in activity for up to 48 h was observed. The activity half-life at 65°C was 48 and 3 h for ASKA and
ADTA, respectively. In the presence of Ca2+ ions, both amylases were highly stable for at least 48 h and had less than a 10% decrease in activity at 70°C. Both enzymes
exhibited similar end-product profiles, and the predominant yield was maltose (69%) from starch hydrolysis. To the best of
our knowledge, most α-amylases that produce high levels of maltose are active at an acidic to neutral pH. This is the first
report of two thermostable, alkalitolerant recombinant α-amylases from Anoxybacillus that produce high levels of maltose and have an atypical protein sequence compared with known α-amylases. 相似文献
92.
Kushwaha GS Pandey N Sinha M Singh SB Kaur P Sharma S Singh TP 《Biochimica et biophysica acta》2012,1824(4):679-691
The ribosome inactivating proteins (RIPs) of type 1 are plant toxins that eliminate adenine base selectively from the single stranded loop of rRNA. We report six crystal structures, type 1 RIP from Momordica balsamina (A), three in complexed states with ribose (B), guanine (C) and adenine (D) and two structures of MbRIP-1 when crystallized with adenosine triphosphate (ATP) (E) and 2'-deoxyadenosine triphosphate (2'-dATP) (F). These were determined at 1.67?, 1.60?, 2.20?, 1.70?, 2.07? and 1.90? resolutions respectively. The structures contained, (A) unbound protein molecule, (B) one protein molecule and one ribose sugar, (C) one protein molecule and one guanine base, (D) one protein molecule and one adenine base, (E) one protein molecule and one ATP-product adenine molecule and (F) one protein molecule and one 2'-dATP-product adenine molecule. Three distinct conformations of the side chain of Tyr70 were observed with (i) χ(1)=-66°and χ(2)=165° in structures (A) and (B); (ii) χ(1)=-95° and χ(2)=70° in structures (C), (D) and (E); and (iii) χ(1)=-163° and χ(2)=87° in structure (F). The conformation of Tyr70 in (F) corresponds to the structure of a conformational intermediate. This is the first structure which demonstrates that the slow conversion of DNA substrates by RIPs can be trapped during crystallization. 相似文献
93.
Gajraj Singh KushwahaNisha Pandey Mau SinhaS. Baskar Singh Punit KaurSujata Sharma Tej P. Singh 《Biochimica et Biophysica Acta - Proteins and Proteomics》2012,1824(4):679-691
The ribosome inactivating proteins (RIPs) of type 1 are plant toxins that eliminate adenine base selectively from the single stranded loop of rRNA. We report six crystal structures, type 1 RIP from Momordica balsamina (A), three in complexed states with ribose (B), guanine (C) and adenine (D) and two structures of MbRIP-1 when crystallized with adenosine triphosphate (ATP) (E) and 2′-deoxyadenosine triphosphate (2′-dATP) (F). These were determined at 1.67 Å, 1.60 Å, 2.20 Å, 1.70 Å, 2.07 Å and 1.90 Å resolutions respectively. The structures contained, (A) unbound protein molecule, (B) one protein molecule and one ribose sugar, (C) one protein molecule and one guanine base, (D) one protein molecule and one adenine base, (E) one protein molecule and one ATP-product adenine molecule and (F) one protein molecule and one 2′-dATP-product adenine molecule. Three distinct conformations of the side chain of Tyr70 were observed with (i) χ1 = − 66°and χ2 = 165° in structures (A) and (B); (ii) χ1 = − 95° and χ2 = 70° in structures (C), (D) and (E); and (iii) χ1 = − 163° and χ2 = 87° in structure (F). The conformation of Tyr70 in (F) corresponds to the structure of a conformational intermediate. This is the first structure which demonstrates that the slow conversion of DNA substrates by RIPs can be trapped during crystallization. 相似文献
94.
Sharma P Singh N Sinha M Sharma S Perbandt M Betzel C Kaur P Srinivasan A Singh TP 《Journal of molecular biology》2008,378(4):923-932
The mammalian peptidoglycan recognition protein-S (PGRP-S) binds to peptidoglycans (PGNs), which are essential components of the cell wall of bacteria. The protein was isolated from the samples of milk obtained from camels with mastitis and purified to homogeneity and crystallized. The crystals belong to orthorhombic space group I222 with a = 87.0 Å, b = 101.7 Å and c = 162.3 Å having four crystallographically independent molecules in the asymmetric unit. The structure has been determined using X-ray crystallographic data and refined to 1.8 Å resolution. Overall, the structures of all the four crystallographically independent molecules are identical. The folding of PGRP-S consists of a central β-sheet with five β-strands, four parallel and one antiparallel, and three α-helices. This protein fold provides two functional sites. The first of these is the PGN-binding site, located on the groove that opens on the surface in the direction opposite to the location of the N terminus. The second site is implicated to be involved in the binding of non-PGN molecules, it also includes putative N-terminal segment residues (1-31) and helix α2 in the extended binding. The structure reveals a novel arrangement of PGRP-S molecules in which two pairs of molecules associate to form two independent dimers. The first dimer is formed by two molecules with N-terminal segments at the interface in which non-PGN binding sites are buried completely, whereas the PGN-binding sites of two participating molecules are fully exposed at the opposite ends of the dimer. In the second dimer, PGN-binding sites are buried at the interface while non-PGN binding sites are fully exposed at the opposite ends of the dimer. This form of dimeric arrangement is unique and seems to be aimed at enhancing the capability of the protein against specific invading bacteria. This mode of functional dimerization enhances efficiency and specificity, and is observed for the first time in the family of PGRP molecules. 相似文献
95.
A genome survey of Moniliophthora perniciosa gives new insights into Witches' Broom Disease of cacao
Jorge MC Mondego Marcelo F Carazzolle Gustavo GL Costa Eduardo F Formighieri Lucas P Parizzi Johana Rincones Carolina Cotomacci Dirce M Carraro Anderson F Cunha Helaine Carrer Ramon O Vidal Raíssa C Estrela Odalys García Daniela PT Thomazella Bruno V de Oliveira Acássia BL Pires Carolina S Maria Rio Marcos Renato R Araújo Marcos H de Moraes Luis AB Castro Karina P Gramacho Marilda S Gonçalves José P Moura Neto Aristóteles Góes Neto Luciana V Barbosa Mark J Guiltinan Bryan A Bailey Lyndel W Meinhardt Julio CM Cascardo Gonçalo AG Pereira 《BMC genomics》2008,9(1):1-25
96.
Michele Guescini Davide Sisti Marco BL Rocchi Laura Stocchi Vilberto Stocchi 《BMC bioinformatics》2008,9(1):326
Background
Real-time PCR analysis is a sensitive DNA quantification technique that has recently gained considerable attention in biotechnology, microbiology and molecular diagnostics. Although, the cycle-threshold (Ct) method is the present "gold standard", it is far from being a standard assay. Uniform reaction efficiency among samples is the most important assumption of this method. Nevertheless, some authors have reported that it may not be correct and a slight PCR efficiency decrease of about 4% could result in an error of up to 400% using the Ct method. This reaction efficiency decrease may be caused by inhibiting agents used during nucleic acid extraction or copurified from the biological sample. 相似文献97.
To establish an adequate model to study the proliferation and differentiation of porcine skeletal muscle in response to bioactive
compounds, a pool of satellite cells was derived from the semimembranosus muscle (SM) of newborn piglets using a Percoll gradient centrifugation. The final yield amounted to 4.1 × 106 cells/g muscle tissue. The percentage of muscle satellite cells has been determined by immunostaining for desmin and subsequent
fluorescence analysis by flow cytometry, which revealed 95% of desmin-positive cells. For proliferation studies, satellite
cell born myoblasts were seeded in gelatin-coated 96-well microplates at about 5 × 103 cells per well. Cells were grown for 1 day in MEMα plus 10% fetal bovine serum (FBS) and 10% horse serum (HS), followed by
2 d cultivation in serum-free growth medium. For differentiation studies, myoblasts were cultured in matrigel-coated 24-well
plates for 4 d with growth medium containing 10% FBS and 10% HS. At 80% confluence, cells were grown for 24 h in medium plus
10% FBS and 1 μM insulin to initiate differentiation. Subsequently, the cells were cultured in serum-free differentiation
medium (SFDM) for 3 d to form myotubes. Cultures reached a maximum fusion rate of approximately 20% after 96 h. By establishing
this culture system, we provide an advanced and appropriate in vitro model to study porcine skeletal muscle cell growth and differentiation including the responses to various bioactive compounds. 相似文献
98.
Using Ultralow Dosages of Electron Acceptor to Reveal the Early Stage Donor–Acceptor Electronic Interactions in Bulk Heterojunction Blends 下载免费PDF全文
Carr Hoi Yi Ho Sin Hang Cheung Ho‐Wa Li Ka Lok Chiu Yuanhang Cheng Hang Yin Mau Hing Chan Franky So Sai‐Wing Tsang Shu Kong So 《Liver Transplantation》2017,7(12)
Tuning the donor–acceptor (D–A) weight ratio is an essential step to optimize the performance of a bulk heterojunction (BHJ) solar cell. The unoptimized regime with a low acceptor concentration is generally unexplored despite it may reveal the early stage electronic D–A interactions. In this study, PTB7:PC71BM is used to examine factors that limit the device performance in unoptimized regime. The key limiting factor is the creation of traps and localized states originated from fullerene molecules. Photothermal deflection spectroscopy is used to quantify the trap density. Starting with pristine PTB7, addition of small concentration of fullerene increases the electron trap density and lowers the electron mobility. When the D–A weight ratio reaches 1:0.1, fullerene percolation occurs. There is an abrupt drop in trap density and simultaneously a six orders of magnitude increase in the electron mobility. Furthermore, the fill factors of the corresponding photovoltaic devices are found to anticorrelate with the trap density. This study reveals that electron trapping is the key limiting factor for unoptimized BHJ solar cells in low fullerene regime. 相似文献
99.
Melissa L. Barker-Haliski Kristina Johnson Peggy Billingsley Jennifer Huff Laura J. Handy Rizvana Khaleel Zhenmei Lu Matthew J. Mau Timothy H. Pruess Carlos Rueda Gerald Saunders Tristan K. Underwood Fabiola Vanegas Misty D. Smith Peter J. West Karen S. Wilcox 《Neurochemical research》2017,42(7):1904-1918
The successful identification of promising investigational therapies for the treatment of epilepsy can be credited to the use of numerous animal models of seizure and epilepsy for over 80 years. In this time, the maximal electroshock test in mice and rats, the subcutaneous pentylenetetrazol test in mice and rats, and more recently the 6 Hz assay in mice, have been utilized as primary models of electrically or chemically-evoked seizures in neurologically intact rodents. In addition, rodent kindling models, in which chronic network hyperexcitability has developed, have been used to identify new agents. It is clear that this traditional screening approach has greatly expanded the number of marketed drugs available to manage the symptomatic seizures associated with epilepsy. In spite of the numerous antiseizure drugs (ASDs) on the market today, the fact remains that nearly 30% of patients are resistant to these currently available medications. To address this unmet medical need, the National Institute of Neurological Disorders and Stroke (NINDS) Epilepsy Therapy Screening Program (ETSP) revised its approach to the early evaluation of investigational agents for the treatment of epilepsy in 2015 to include a focus on preclinical approaches to model pharmacoresistant seizures. This present report highlights the in vivo and in vitro findings associated with the initial pharmacological validation of this testing approach using a number of mechanistically diverse, commercially available antiseizure drugs, as well as several probe compounds that are of potential mechanistic interest to the clinical management of epilepsy. 相似文献
100.
Humans play major roles in shaping and transforming the ecology of Earth. Unlike natural drivers of ecosystem change, which are erratic and unpredictable, human intervention in ecosystems generally involves planning and management, but often results in detrimental outcomes. Using model studies and aerial-image analysis, we argue that the design of a successful human intervention form calls for the identification of the self-organization modes that drive ecosystem change, and for studying their dynamics. We demonstrate this approach with two examples: grazing management in drought-prone ecosystems, and rehabilitation of degraded vegetation by water harvesting. We show that grazing can increase the resilience to droughts, rather than imposing an additional stress, if managed in a spatially non-uniform manner, and that fragmental restoration along contour bunds is more resilient than the common practice of continuous restoration in vegetation stripes. We conclude by discussing the need for additional studies of self-organization modes and their dynamics. 相似文献