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311.
Oncogenic Raf-1 disrupts epithelial tight junctions via downregulation of occludin 总被引:11,自引:0,他引:11
Occludin is an integral membrane protein of the epithelial cell tight junction (TJ). Its potential role in coordinating structural and functional events of TJ formation has been suggested recently. Using a rat salivary gland epithelial cell line (Pa-4) as a model system, we have demonstrated that occludin not only is a critical component of functional TJs but also controls the phenotypic changes associated with epithelium oncogenesis. Transfection of an oncogenic Raf-1 into Pa-4 cells resulted in a complete loss of TJ function and the acquisition of a stratified phenotype that lacked cell-cell contact growth control. The expression of occludin and claudin-1 was downregulated, and the distribution patterns of ZO-1 and E-cadherin were altered. Introduction of the human occludin gene into Raf-1-activated Pa-4 cells resulted in reacquisition of a monolayer phenotype and the formation of functionally intact TJs. In addition, the presence of exogenous occludin protein led to a recovery in claudin-1 protein level, relocation of the zonula occludens 1 protein (ZO-1) to the TJ, and redistribution of E-cadherin to the lateral membrane. Furthermore, the expression of occludin inhibited anchorage-independent growth of Raf-1-activated Pa-4 cells in soft agarose. Thus, occludin may act as a pivotal signaling molecule in oncogenic Raf- 1-induced disruption of TJs, and regulates phenotypic changes associated with epithelial cell transformation. 相似文献
312.
In this study, floral spray and floral dip were used to replace the vacuum step in the Agrobacterium-mediated transformation of a superoxide dismutase (SOD) gene into Arabidopsis. The transgene was constructed by using a CaMV 35S promoter to drive a rice cytosolic CuZnSOD coding sequence in Arabidopsis. The transgene construct was developed in binary vectors and mobilized into Agrobacterium. When Arabidopsis plants started to initiate flower buds, the primary inflorescence shoots were removed and then transformed by floral spray or floral dip. More than 300 transgenic plants were generated to assess the feasibility of floral spray used in the in planta transformation. The result indicates that the floral spray method of Agrobacterium can achieve rates of in planta transformation comparable to the vacuum-infiltration and floral dip methods. The floral spray method opens up the possibility of in planta transformation of plant species which are too large for dipping or vacuum infiltration. 相似文献
313.
Krause K Maier RM Kofer W Krupinska K Herrmann RG 《Molecular & general genetics : MGG》2000,263(6):1022-1030
314.
One of the photosystem II reaction center proteins, D1, is encoded by the psbA gene and is synthesized as a precursor form with a carboxyl-terminal extension that is subsequently cleaved between Ala-344 and Ser-345. We have generated three psbA transformants of the green alga Chlamydomonas reinhardtii in which Ala-344 or Ser-345 have been substituted with Pro or Glu (A344P, S345E, and S345P) to understand the effects of the amino acid substitutions on the processing of the precursor D1. S345E grew photoautotrophically and showed PSII activity like the wild type. However, A344P and S345P were unable to grow photoautotrophically and were significantly photosensitive. A344P was deficient in the processing of precursor D1 and in oxygen-evolving activity, but assembled photosystem II complex capable of charge separation. In contrast, both precursor and mature forms of D1 accumulated in S345P cells from the logarithmic phase and the cells evolved oxygen at 18% of wild-type level. However, S345P cells from the stationary phase contained mostly the mature D1 and showed a twofold increase in oxygen-evolving activity. The rate of processing of the accumulated pD1 was estimated to be about 100 times slower than in the wild type. It is therefore concluded that the functional oxygen-evolving complex is assembled when the precursor D1 is processed, albeit at a very low rate. These results suggest the functional significance of the amino acid residues at the processing site of the precursor D1. 相似文献
315.
Gil A. Enríquez-Obregón Roberto I. Vázquez-Padrón Dmitri L. Prieto-Samsonov Gustavo A. De la Riva Guillermo Selman-Housein 《Planta》1998,206(1):20-27
The presence of undesirable plants in sugarcane (Saccharum officinarum L.) plantations reduces crop yields. Using genetic engineering as a complement for traditional breeding methods it is possible
to introduce herbicide-resistant traits into Saccharum germplasm. Transgenic sugarcane plants resistant to phosphinothricine (PPT), the active compound of the commercial herbicide
BASTA were generated by Agrobacterium tumefaciens-mediated transformation. Meristematic sections of sugarcane were treated with anti-necrotic compounds to minimize oxidative
bursts and used as explants. Four transformation protocols were assessed and the transformation frequencies reached 10–35%.
The regeneration rate was high and did not appear to be affected by the transformation procedure. Southern blot analysis of
several transformed plants indicated the integration per genome of one or two intact copies of the bar gene which encodes PPT acetyltransferase and confers resistance to BASTA. The levels of BASTA resistance were evaluated under
greenhouse and small-plot conditions.
Received: 8 November 1997 / Accepted: 22 November 1997 相似文献
316.
H. Tsugawa Y. Otsuki M. Suzuki 《TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik》1998,97(7):1019-1026
A very simple and efficient transformation system for rice was established using a synthetic polycationic amino polymer (polycation).
Improvements in the culture conditions, especially filtration of the suspension cells before and after protoplast culture,
greatly contributed to a large yield of high-quality protoplasts and an increased ability of the cells to regenerate. Transformation
parameters, such as the ratio of DNA and polycation concentrations, preincubation of the DNA and polycation prior to DNA transfer,
and precentrifugation and resuspension of protoplasts before DNA transfer, were analyzed. Fertile transgenic plants containing
the bar gene were selected and shown to demonstrate resistance against high concentrations of bialaphos. Southern blot analysis showed
four to nine bands representing the bar gene in polycation-mediated transgenic rice plants compared with two to three bands in electroporation-mediated transgenic
rice plants. The regeneration efficiency of the polycation-mediated method was compared to that of the electroporation-mediated
method; while the polycation-mediated method tended to show a relatively lower regeneration rate, regenerants showed a normal
phenotype.
Received: 26 February 1998 / Accepted: 15 May 1998 相似文献
317.
Plastid transformation in Arabidopsis thaliana 总被引:33,自引:0,他引:33
Plastid transformation is reported in Arabidopsis thaliana following biolistic delivery of transforming DNA into leaf cells. Transforming plasmid pGS31A carries a spectinomycin resistance
(aadA) gene flanked by plastid DNA sequences to target its insertion between trnV and the rps12/7 operon. Integration of aadA by two homologous recombination events via the flanking ptDNA sequences and selective amplification of the transplastomes
on spectinomycin medium yielded resistant cell lines and regenerated plants in which the plastid genome copies have been uniformly
altered. The efficiency of plastid transformation was low: 2 in 201 bombarded leaf samples. None of the 98 plants regenerated
from the two lines were fertile.
Received: 13 February 1998 / Revision received: 24 April 1998 / Accepted: 5 June 1998 相似文献
318.
Transgenic Japanese lawngrass (Zoysia japonica Steud.) plants regenerated from protoplasts 总被引:14,自引:0,他引:14
Transgenic Japanese lawngrass (Zoysia japonica Steud.) plants were generated by means of polyethylene glycol (PEG)-mediated direct gene transfer into protoplasts. The plasmid
pBC1 was used to deliver the hygromycin phosphotransferase (hph) and β-glucuronidase (gus) genes into protoplasts. Selection with a high concentration (400 mg/l) of hygromycin yielded a number of resistant calli
and about 400 plants were generated. Polymerase chain reaction (PCR) and Southern hybridization analyses revealed that all
of then plants tested contained introduced genes. The gus gene regulated by the maize alcohol dehydrogenase-1 (Adh 1) promoter was expressed in the leaves and roots of transgenic Japanese lawngrass plants.
Received: 13 December 1996 / Revision received: 9 June 1997 / Accepted: 2 September 1997 相似文献
319.
Daniel K.Y. Solaiman 《Biotechnology Techniques》1998,12(11):829-832
An electroporation procedure for the transformation of Pseudomonas oleovorans was developed using a model plasmid, pCN51. The optimal electrotransformation was achieved with cells harvested at 45 to 60 min of growth and concentrated to a cell density of 5 OD600nm, plasmid concentration of 6 g per 100 l of cell suspension, and a 0.1-cm gap-width cuvette. Electroporation was performed at the settings of 250 , 25F and 2.5 kV. Transformation yields in the order of 103 colony-forming-unit per electroporation sample were obtained. This is a first report of the electroporation of the commercially valuable bacterium Ps. oleovorans. © Rapid Science Ltd. 1998 相似文献
320.
用BamH1和CIP处理pCB20质粒,Sau3Al部分降解枯草杆菌抗阿霉素突变株染色体DNA,T4连接酶连接载体和DNA片断,构成表达文库。用这个文库分别转化枯草杆菌BD224菌株感受态细胞和原生质体。 相似文献