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171.
Active Subtilisin-Like Protease from a Hyperthermophilic Archaeon in a Form with a Putative Prosequence 总被引:6,自引:2,他引:4
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Yuji Kannan Yuichi Koga Yohei Inoue Mitsuru Haruki Masahiro Takagi Tadayuki Imanaka Masaaki Morikawa Shigenori Kanaya 《Applied microbiology》2001,67(6):2445-2452
The gene encoding subtilisin-like protease T. kodakaraensis subtilisin was cloned from a hyperthermophilic archaeon Thermococcus kodakaraensis KOD1. T. kodakaraensis subtilisin is a member of the subtilisin family and composed of 422 amino acid residues with a molecular weight of 43,783. It consists of a putative presequence, prosequence, and catalytic domain. Like bacterial subtilisins, T. kodakaraensis subtilisin was overproduced in Escherichia coli in a form with a putative prosequence in inclusion bodies, solubilized in the presence of 8 M urea, and refolded and converted to an active molecule. However, unlike bacterial subtilisins, in which the prosequence was removed from the catalytic domain by autoprocessing upon refolding, T. kodakaraensis subtilisin was refolded in a form with a putative prosequence. This refolded protein of recombinant T. kodakaraensis subtilisin which is composed of 398 amino acid residues (Gly−82 to Gly316), was purified to give a single band on a sodium dodecyl sulfate (SDS)-polyacrylamide gel and characterized for biochemical and enzymatic properties. The good agreement of the molecular weights estimated by SDS-polyacrylamide gel electrophoresis (44,000) and gel filtration (40,000) suggests that T. kodakaraensis subtilisin exists in a monomeric form. T. kodakaraensis subtilisin hydrolyzed the synthetic substrate N-succinyl-Ala-Ala-Pro-Phe-p-nitroanilide only in the presence of the Ca2+ ion with an optimal pH and temperature of pH 9.5 and 80°C. Like bacterial subtilisins, it showed a broad substrate specificity, with a preference for aromatic or large nonpolar P1 substrate residues. However, it was much more stable than bacterial subtilisins against heat inactivation and lost activity with half-lives of >60 min at 80°C, 20 min at 90°C, and 7 min at 100°C. 相似文献
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Kazutomo Haraguchi Mitsuru Yoshida Tomomi Yamanaka Kenichi Ohtsubo 《Carbohydrate polymers》2003,53(4):501-505
An inulin fructotransferase (DFA I-producing) [EC 2.4.1.200] from Arthrobacter pascens a62-1 was purified and the properties of the enzyme were investigated. The enzyme was purified from culture supernatant of the microorganism 58.5 fold with a yield of 8.32% using Super Q Toyopearl chromatography and butyl Toyopearl chromatography. It showed maximum activity at pH 5.5 and 45 °C and was stable up to 75 °C. This heat stability was highest in the inulin fructotransferases (DFA I-producing) reported until now. The molecular mass of the enzyme was estimated to be 37,000 by SDS-PAGE and 60,000 by gel filtration, and was considered to be a dimer. The N-terminal amino acid sequence (20 amino acid residues) was determined as Ala-Asn-Thr-Val-Tyr-Asp-Val-Thr-Thr-Trp-Ser-Gly-Ala-Thr-Ile-Ser-Pro-Tyr-Val-Asp. 相似文献
174.
The Frequent 5,10-Methylenetetrahydrofolate Reductase C677T Polymorphism Is Associated with a Common Haplotype in Whites, Japanese, and Africans 总被引:9,自引:0,他引:9
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Nurit Rosenberg Mitsuru Murata Yasuo Ikeda Ohene Opare-Sem Ariella Zivelin Eli Geffen Uri Seligsohn 《American journal of human genetics》2002,70(3):758-762
The common 5,10-methylenetetrahydrofolate reductase (MTHFR) C677T polymorphism causes decreased activity of this enzyme and can be associated with mild-to-moderate hyperhomocysteinemia in homozygotes, particularly when there is folic acid deficiency, as well as with vascular dementia, arterial thrombosis, venous thrombosis, neural-tube defects, and fetal loss. When folic acid intake is sufficient, homozygotes for MTHFR 677T appear to be protected against colon cancer and acute lymphatic leukemia, and fetuses bearing this genotype have an augmented survival. The distribution of MTHFR 677T is worldwide, but its frequency in different populations varies extensively. In the present study, we addressed the question of whether the MTHFR 677T alteration has an ancestral origin or has occurred repeatedly. We analyzed the frequency distribution of the previously described polymorphism A1298C in exon 7 and of three intronic dimorphisms, in white Israelis (Jews and Arabs), Japanese, and Ghanaian Africans. The 677T allele was, remarkably, associated with one haplotype, G-T-A-C, in white and Japanese homozygotes. Among the Africans, analysis of maximum likelihood also disclosed an association with the G-T-A-C haplotype, although none of the 174 subjects examined was homozygous for MTHFR 677T. These results suggest that the MTHFR 677T alteration occurred on a founder haplotype that may have had a selective advantage. 相似文献
175.
Toshiro Nishimura Shigekazu Teranishi Atsuhiro Kawashima Tadashi Ishimaru Takaki Miwa Mitsuru Furukawa 《Chemical senses》2002,27(1):13-21
Systemic or topical application of glucocorticoid is the treatment of choice for olfactory disturbance. Recently, Na(+)/K(+) ATPase and glucocorticoid receptor immunoreactivity in the olfactory mucosa was reported. To elucidate a glucocorticoid action on Na(+)/K(+) ATPase production, an animal model was produced by an intra-nasal application of 5% ZnSO(4) solution to Wistar rats. Dexamethasone was injected i.p. (0.01 mg/100 g) for 14 days after the insult. Histologically, the regeneration process was completed on day 14 in both dexamethasone- and saline-injected control rats. We used a quantitative polymerase chain reaction (PCR) method to evaluate mRNA production of Na(+)/K(+) ATPase and glucocorticoid receptor. In dexamethasone-injected rats, up-regulation of glucocorticoid receptor mRNA (95% more than control rats, P = 0.00068, unpaired t-test) and of Na(+)/K(+) ATPase mRNA expression (76% more than control rats, P = 0.0042) was observed on day 14. The increased Na(+)/K(+) ATPase expression in the regenerated olfactory mucosa is thought to be beneficial for an active uptake of K(+), which is released during excitation, around olfactory neurons and for the transepithelial absorption of Na(+) from olfactory mucus. Dexamethasone may thus contribute to the recovery of function after the morphological regeneration in part, at least, through its receptor by regulation of the ionic concentration in the olfactory mucosal microenvironment. 相似文献
176.
Structure of human holocarboxylase synthetase gene and mutation spectrum of holocarboxylase synthetase deficiency 总被引:4,自引:0,他引:4
Xue Yang Yoko Aoki Xue Li Osamu Sakamoto Masahiro Hiratsuka Shigeo Kure Sepidh Taheri Ernst Christensen Koji Inui Mitsuru Kubota Miki Ohira Misao Ohki Jun Kudoh Kazuhiko Kawasaki Kazunori Shibuya Ai Shintani Shuichi Asakawa Shinsei Minoshima Nobuyoshi Shimizu Kuniaki Narisawa Yoichi Matsubara Yoichi Suzuki 《Human genetics》2001,109(5):526-534
Holocarboxylase synthetase (HLCS) is an enzyme that catalyzes the incorporation of biotin into apo-carboxylases, and its deficiency causes biotin-responsive multiple carboxylase deficiency. The reported sequences of cDNA for human HLCS from liver, lymphocyte, and KG-1 myeloid cell lines differ at their 5' regions. To elucidate variations of the human HLCS mRNA and longer 5' cDNA ends, we performed screening of the human liver cDNA library and rapid amplification of the cDNA ends (RACE). Our results suggest the existence of three types of HLCS mRNA that start at different exons. The first type starts at exon 1, and the second type starts at exon 3, and both are found in various human tissues. The third type, corresponding to the cDNA from the KG-1 cell, starts at exon 2 of the HLCS gene. Various splicing patterns from exons 3-6 were also observed. None of the variations of cDNA found created a new initiation codon. Mutation screening from exons 6-14, therefore, was sufficient to detect amino acid changes in HLCS in patients. Our direct sequencing strategy for screening mutations in the HLCS gene revealed mutations in five Japanese patients and seven non-Japanese patients. Our analyses involving 12 Japanese and 13 non-Japanese patients and studies by others indicate that (1) there is no panethnically prevalent mutation; (2) the Arg508Trp, Gly581Ser, and Val550Met mutations are found in both Japanese and non-Japanese populations; (3) the IVS10+5G-->A mutation is predominant and probably a founder mutation in European patients; (4) the 655-656insA, Leu237Pro, and 780delG mutations are unique in Japanese patients; (5) the spectrum of the mutations in the HLCS gene may vary substantially among different ethnic groups. 相似文献
177.
Masato Nikaido Fumio Matsuno Hideaki Abe Mitsuru Shimamura Healy Hamilton Hisashi Matsubayashi Norihiro Okada 《Mammalian genome》2001,12(12):909-915
Short interspersed repetitive elements (SINEs) are a kind of retroposons dispersed among the eukaryotic genomes. Previously,
we isolated and characterized a new SINE family, named CHR-2, members of which are distributed in the genomes of cetaceans,
hippopotamuses, and ruminants. We analyzed systematically more than a hundred members of the CHR-2 SINEs, which were isolated
from the genomes of cetaceans and cow, together with the additional data available in the DNA databases, and showed that these
SINEs are divided into at least five distinct subfamilies that share diagnostic nucleotides and/or deletions. A hybridization
analysis clearly demonstrated that, among these five subfamilies, two subfamilies, named CD and CDO, are specific to cetaceans
and toothed whales, respectively. We reconstruct the evolutionary history of the CHR-2 SINEs during evolution of cetartiodactyl
genomes.
Received: 13 June 2001 / Accepted: 12 July 2001 相似文献
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