全文获取类型
收费全文 | 1497篇 |
免费 | 205篇 |
出版年
2021年 | 15篇 |
2016年 | 17篇 |
2015年 | 27篇 |
2014年 | 33篇 |
2013年 | 46篇 |
2012年 | 84篇 |
2011年 | 59篇 |
2010年 | 51篇 |
2009年 | 37篇 |
2008年 | 59篇 |
2007年 | 57篇 |
2006年 | 70篇 |
2005年 | 66篇 |
2004年 | 62篇 |
2003年 | 68篇 |
2002年 | 58篇 |
2001年 | 57篇 |
2000年 | 58篇 |
1999年 | 53篇 |
1998年 | 21篇 |
1997年 | 12篇 |
1996年 | 18篇 |
1995年 | 10篇 |
1994年 | 14篇 |
1993年 | 21篇 |
1992年 | 27篇 |
1991年 | 35篇 |
1990年 | 33篇 |
1989年 | 31篇 |
1988年 | 27篇 |
1987年 | 24篇 |
1986年 | 35篇 |
1985年 | 31篇 |
1984年 | 26篇 |
1983年 | 19篇 |
1982年 | 34篇 |
1981年 | 13篇 |
1980年 | 21篇 |
1979年 | 15篇 |
1978年 | 18篇 |
1977年 | 14篇 |
1975年 | 13篇 |
1974年 | 15篇 |
1973年 | 11篇 |
1972年 | 18篇 |
1971年 | 13篇 |
1970年 | 10篇 |
1969年 | 16篇 |
1967年 | 9篇 |
1966年 | 7篇 |
排序方式: 共有1702条查询结果,搜索用时 31 毫秒
1.
Eleanor R. Watkins Bridget S. Penman José Louren?o Caroline O. Buckee Martin C. J. Maiden Sunetra Gupta 《PLoS pathogens》2015,11(7)
The bacterial pathogen, Streptococcus pneumoniae (the pneumococcus), is a leading cause of life-threatening illness and death worldwide. Available conjugate vaccines target only a small subset (up to 13) of >90 known capsular serotypes of S. pneumoniae and, since their introduction, increases in non-vaccine serotypes have been recorded in several countries: a phenomenon termed Vaccine Induced Serotype Replacement (VISR). Here, using a combination of mathematical modelling and whole genome analysis, we show that targeting particular serotypes through vaccination can also cause their metabolic and virulence-associated components to transfer through recombination to non-vaccine serotypes: a phenomenon we term Vaccine-Induced Metabolic Shift (VIMS). Our results provide a novel explanation for changes observed in the population structure of the pneumococcus following vaccination, and have important implications for strain-targeted vaccination in a range of infectious disease systems. 相似文献
2.
Gene expression in visceral endoderm: a comparison of mutant and wild- type F9 embryonal carcinoma cell differentiation 下载免费PDF全文
We have examined the abundance and cell specificity of several mRNAs that are regulated during the retinoic acid (RA)-induced differentiation of F9 embryonal carcinoma cells to visceral endoderm. The experiments confirmed the multistep nature of this process by demonstrating the expression of the ERA-1/Hox 1.6 message within 6 h after RA addition; the expression of messages specific for the extracellular matrix proteins laminin B1 and B2, and collagen IV(alpha 1) between days 4 and 12; and the expression of two visceral endoderm markers, alpha-fetoprotein (AFP) and H19, by days 8-15. In situ hybridization experiments revealed that the collagen IV(alpha 1) mRNA is restricted to the outer cell layer of F9 cell aggregates regardless of the presence or absence of RA. Laminin B1 and B2 mRNAs are concentrated in the outer cell layer of RA-treated aggregates although significant levels of message are also observed within the interior cells of the aggregates. Unexpectedly, AFP mRNA is detectable in only a subset of the outer cells of F9 cell aggregates grown 15 d in the presence of RA. The results obtained from wild-type F9 cells were compared with those from a mutant F9 cell line, RA-5-1, which was previously shown to synthesize collagen IV containing six- to ninefold less 4-hydroxyproline than that in wild-type F9 cells. RA-5-1 cells exhibit four- to sixfold less of the mRNAs encoding two visceral endoderm proteins, AFP and H19, than wild-type F9 cells after RA treatment of RA-5-1 aggregates. RA-5-1 cells, however, do exhibit an RA-associated increase in the level of ERA-1/Hox 1.6 mRNA within 6 h after adding RA. Although the collagen IV protein level is similar in wild-type F9 and RA-5-1 aggregates, the collagen IV(alpha 1) message level is 6-20-fold greater in aggregates of mutant cells than in aggregates of wild-type cells. Moreover, in situ hybridizations showed that this message is evenly distributed throughout the RA-5-1 aggregates rather than restricted to the outer cell layers as it is in wild-type F9 aggregates. These results suggest that abnormal collagen IV expression and localization are associated with decreased expression of the visceral endoderm markers, AFP and H19, in RA-5-1 cell aggregates. 相似文献
3.
Detection of a specific isoform of alpha-actinin with antisera directed against dystrophin 总被引:9,自引:5,他引:4 下载免费PDF全文
We have characterized a protein immunologically related to dystrophin, the protein product of the Duchenne muscular dystrophy gene. We identify this related protein as a fast-twitch glycolytic isoform (mouse extensor digitorum longus-specific) of myofibrillar alpha-actinin. This specific isoform of alpha-actinin exhibits a more restricted pattern of expression in skeletal muscle than fast-twitch-specific isoforms of both myosin and Ca2+-ATPase. Our results provide evidence that dystrophin and myofibrillar alpha-actinin are related proteins, reinforcing the previous data concerning the sequence homologies noted between nonmuscle cytoskeletal alpha-actinin and dystrophin. In addition, we describe the first antisera directed against a specific myofibrillar skeletal muscle isoform of alpha-actinin. 相似文献
4.
Skewed X inactivation in a female MZ twin results in Duchenne muscular dystrophy. 总被引:16,自引:7,他引:9 下载免费PDF全文
C S Richards S C Watkins E P Hoffman N R Schneider I W Milsark K S Katz J D Cook L M Kunkel J M Cortada 《American journal of human genetics》1990,46(4):672-681
One of female MZ twins presented with muscular dystrophy. Physical examination, creatine phosphokinase levels, and muscle biopsy were consistent with Duchenne muscular dystrophy (DMD). However, because of her sex she was diagnosed as having limb-girdle muscular dystrophy. With cDNA probes to the DMD gene, a gene deletion was detected in the twins and their mother. The de novo mutation which arose in the mother was shown by novel junction fragments generated by HindIII, PstI, or TaqI when probed with cDNA8. Additional evidence of a large gene deletion was given by novel SfiI junction fragments detected by probes p20, J-Bir, and J-66 on pulsed-field gel electrophoresis (PFGE). Immunoblot analysis of muscle from the affected twin showed dystrophin of normal size but of reduced amount. Immunofluorescent visualization of dystrophin revealed foci of dystrophin-positive fibers adjacent to foci of dystrophin-negative fibers. These data indicate that the affected twin is a manifesting carrier of an abnormal DMD gene, her myopathy being a direct result of underexpression of dystrophin. Cytogenetic analysis revealed normal karyotypes, eliminating the possibility of a translocation affecting DMD gene function. Both linkage analysis and DNA fingerprint analysis revealed that each twin has two different X chromosomes, eliminating the possibility of uniparental disomy as a mechanism for DMD expression. On the basis of methylation differences of the paternal and maternal X chromosomes in these MZ twins, we propose uneven lyonization (X chromosome inactivation) as the underlying mechanism for disease expression in the affected female. 相似文献
5.
A genetic linkage map of human chromosome 21: analysis of recombination as a function of sex and age. 总被引:10,自引:6,他引:4 下载免费PDF全文
R E Tanzi P C Watkins G D Stewart N S Wexler J F Gusella J L Haines 《American journal of human genetics》1992,50(3):551-558
A genetic linkage map of human chromosome 21 has been constructed using 22 anonymous DNA markers and five complementary DNAs (cDNAs) encoding the amyloid beta protein precursor (APP), superoxide dismutase 1 (SOD1), the ets-2 proto-oncogene (ETS2), the estrogen inducible breast cancer locus (BCEI), and the leukocyte antigen, CD18 (CD18). Segregation of RFLPs detected by these DNA markers was traced in the Venezuelan Reference Pedigree (VRP). A comprehensive genetic linkage map consisting of the 27 DNA markers spans 102 cM on the long arm of chromosome 21. We have confirmed our initial findings of a dramatically increased rate of recombination at the telomere in both females and males and of significantly higher recombination in females in the pericentromeric region. By comparing patterns of recombination in specific regions of chromosome 21 with regard to both parental sex and age, we have now identified a statistically significant downward trend in the frequency of crossovers in the most telomeric portion of chromosome 21 with increasing maternal age. A less significant decrease in recombination with increasing maternal age was observed in the pericentromeric region of the chromosome. These results may help in ultimately understanding the physical relationship between recombination and nondisjunction in the occurrence of trisomy 21. 相似文献
6.
The location of CTP:phosphocholine cytidylyltransferase in Chinese hamster ovary (CHO) cells made deficient in phosphatidylcholine was determined by immunofluorescence techniques. A rabbit polyclonal antibody was raised against a synthetic peptide corresponding to the amino-terminal 17 amino acid residues of rat liver cytidylyltransferase. The antibody recognized both native and denatured cytidylyltransferase from both rat liver and CHO cells. CHO cells were treated with phospholipase C to alter the lipid composition of the plasma membrane and to elicit translocation of cytidylyltransferase from the less active soluble pool to an activated membrane fraction. Visualization of cytidylyltransferase by indirect immunofluorescence revealed staining of the nuclear envelope in phospholipase C-treated cells but not in untreated cells. CHO cells were also starved for choline and supplemented with a choline analogue to provide an alternative technique of rendering the cells phosphatidylcholine-deficient. Although this treatment should affect different cellular membranes than those affected by phospholipase C treatment, cytidylyltransferase still translocated to the nuclear envelope, as shown by indirect immunofluorescence. These results indicate that activated, membrane-bound cytidylyltransferase is associated with the nuclear membrane and suggest that the nuclear membrane may be a site of de novo phosphatidylcholine synthesis. 相似文献
7.
8.
9.
A “parallel plate” electrostatic model for bimolecular rate constants applied to electron transfer proteins 下载免费PDF全文
J. Abra Watkins Michael A. Cusanovich Terrance E. Meyer Gordon Tollin 《Protein science : a publication of the Protein Society》1994,3(11):2104-2114
A “parallel plate” model describing the electrostatic potential energy of protein-protein interactions is presented that provides an analytical representation of the effect of ionic strength on a bimolecular rate constant. The model takes into account the asymmetric distribution of charge on the surface of the protein and localized charges at the site of electron transfer that are modeled as elements of a parallel plate condenser. Both monopolar and dipolar interactions are included. Examples of simple (monophasic) and complex (biphasic) ionic strength dependencies obtained from experiments with several electron transfer protein systems are presented, all of which can be accommodated by the model. The simple cases do not require the use of both monopolar and dipolar terms (i.e., they can be fit well by either alone). The biphasic dependencies can be fit only by using dipolar and monopolar terms of opposite sign, which is physically unreasonable for the molecules considered. Alternatively, the high ionic strength portion of the complex dependencies can be fit using either the monopolar term alone or the complete equation; this assumes a model in which such behavior is a consequence of electron transfer mechanisms involving changes in orientation or site of reaction as the ionic strength is varied. Based on these analyses, we conclude that the principal applications of the model presented here are to provide information about the structural properties of intermediate electron transfer complexes and to quantify comparisons between related proteins or site-specific mutants. We also conclude that the relative contributions of monopolar and dipolar effects to protein electron transfer kinetics cannot be evaluated from experimental data by present approximations. 相似文献
10.
Jung Joo Hong Matthew R. Reynolds Teresa L. Mattila Aaron Hage David I. Watkins Christopher J. Miller Pamela J. Skinner 《PloS one》2009,4(1)
CD8 T cells play an important role in controlling viral infections. We investigated the in situ localization of simian immunodeficiency virus (SIV)-specific T cells in lymph and genital tissues from SIV-infected macaques using MHC-class I tetramers. The majority of tetramer-binding cells localized in T cell zones and were CD8+. Curiously, small subpopulations of tetramer-binding cells that had little to no surface CD8 were detected in situ both early and late post-infection, and in both vaginally and rectally inoculated macaques. These tetramer+CD8low/− cells were more often localized in apparent B cell follicles relative to T cell zones and more often found near or within the genital epithelium than the submucosa. Cells analyzed by flow cytometry showed similar populations of cells. Further immunohistological characterization revealed small populations of tetramer+CD20− cells inside B cell follicles and that tetramer+ cells did not stain with γδ-TCR nor CD4 antibodies. Negative control tetramer staining indicated that tetramer+CD8low/− cells were not likely NK cells non-specifically binding to MHC tetramers. These findings have important implications for SIV-specific and other antigen-specific T cell function in these specific tissue locations, and suggest a model in which antigen-specific CD8+ T cells down modulate CD8 upon entering B cell follicles or the epithelial layer of tissues, or alternatively a model in which only antigen-specific CD8 T cells that down-modulate CD8 can enter B cell follicles or the epithelium. 相似文献