全文获取类型
收费全文 | 1718篇 |
免费 | 53篇 |
国内免费 | 171篇 |
出版年
2024年 | 1篇 |
2023年 | 9篇 |
2022年 | 8篇 |
2021年 | 11篇 |
2020年 | 18篇 |
2019年 | 22篇 |
2018年 | 20篇 |
2017年 | 21篇 |
2016年 | 21篇 |
2015年 | 28篇 |
2014年 | 44篇 |
2013年 | 78篇 |
2012年 | 48篇 |
2011年 | 75篇 |
2010年 | 61篇 |
2009年 | 65篇 |
2008年 | 70篇 |
2007年 | 68篇 |
2006年 | 70篇 |
2005年 | 62篇 |
2004年 | 68篇 |
2003年 | 78篇 |
2002年 | 78篇 |
2001年 | 60篇 |
2000年 | 69篇 |
1999年 | 42篇 |
1998年 | 61篇 |
1997年 | 30篇 |
1996年 | 29篇 |
1995年 | 46篇 |
1994年 | 47篇 |
1993年 | 42篇 |
1992年 | 49篇 |
1991年 | 36篇 |
1990年 | 29篇 |
1989年 | 37篇 |
1988年 | 36篇 |
1987年 | 40篇 |
1986年 | 45篇 |
1985年 | 53篇 |
1984年 | 50篇 |
1983年 | 31篇 |
1982年 | 31篇 |
1981年 | 17篇 |
1980年 | 14篇 |
1979年 | 16篇 |
1978年 | 6篇 |
1976年 | 2篇 |
排序方式: 共有1942条查询结果,搜索用时 15 毫秒
991.
The Drosophila Schneider S2 (S2) Expression System enables expression of recombinant proteins constitutively, as well as inductively. This
system can establish both transient and stable transformants with various selection markers. The generation of stable cell
lines for increased expression or large scale expression of the desired protein is currently accomplished by cotransfection
of both the expression and selection vectors. The selection vectors, pCoHYGRO and pCoBLAST, are commercially available using
hygromycin-B and blasticidin S, respectively. Recently, we generated a plasmid, pCoPURO, for selection of transfected S2 cells
using puromycin, which allows significant acceleration of the selection time. Although co-transfection of the selection marker
with the plasmid for heterologous protein expression is functional in stable expression at short culture periods, the expression
levels of stable transformants are continuously decreased during long culture times. To overcome this limitation, we generated
pMT-PURO, a new plasmid that contains both the expression cassette and puromycin selection marker in a single plasmid. This
system allows rapid selection and maintenance of the transformed S2 lines for extended culture periods. 相似文献
992.
CTX-M- and AmpC-type beta-lactamases comprise the two most rapidly growing populations among the extended-spectrum cephalosporinases. The evolution and dissemination of resistance genes encoding these enzymes occur mostly through the transmission of plasmids. The high prevalence of clinical isolates of Enterobacteriaceae producing the plasmid-mediated extended-spectrum cephalosporinases resembles an epidemic of plasmids, and has generated serious therapeutic problems. This review describes the emergence and worldwide spread of various classes of plasmid-mediated extended-spectrum cephalosporinases in Salmonella and other Enterobacteriaceae, the transfer mechanism of the plasmids, detection methods, and therapeutic choices. 相似文献
993.
994.
Aims: The calcium chloride chemical transformation of Escherichia coli is still the most widley used cloning method in small laboratories. Therefore, any practicable improvement in its transformation efficiency seems to be of general interest. Methods and Results: We found that giving calcium chloride competent cells a 1 min microwave pulse at the lowest power setting (180 W), instead of the classic 1–2 min 42°C heat‐shock step, increases the transformation efficiency around threefold (3.3 ± 0.5). Moreover, when both treatments were given in a 2‐min 42°C ? 5 min on ice ? 1 min microwave pluse sequence, an additional improvement of 1.6 was obtained, resulting in an overall increase in efficiency of approximately 5.3‐fold compared to classical heat shock. Conclusions: This transformation method significantly improves the classical heat shock treatment. Significance and Impact of the Study: This method might be useful to those laboratories that cannot afford an electroporation apparatus. 相似文献
995.
A systematic analysis of the inheritance of D plasmids of the IncP-9 group (α-, β-, γ-, δ-, ?-, ζ-, η-, and θ-subgroups), IncP-7, as well as of those of undefined systematic affiliation in the cells of homologous (Pseudomonas putida) and heterologous (Escherichia coli) hosts was performed for the first time. For this purpose, mini-Tn5 transposons determining resistance to kanamycin (or streptomycin) were introduced into all the D plasmids under study. It has been established that all IncP-9 plasmids can be transmitted to the cells of a heterologous host E. coli (with the exception of plasmid pSVS15 from gq-subgroup). IncP-7 plasmids and those of undefined systematic affiliation do not possess this property and can be transmitted and stably inherited only in P. putida. The distinctive feature of most IncP-9 plasmids (α-, β-, δ-, ?-, and ζ-subgroups) is strict dependence of their inheritance on the temperature factor. At 37°C, the plasmids of δ-, ζ-, and θ-subgroups are unstable in P. putida cells, while in E. coli nearly all plasmids of this systematic group are unstable. The exceptions are the plasmids of η- and γ-subgroups. Inheritance of these plasmids does not depend on temperature. At 28°C and 37°C, the η plasmid is not maintained stably (inheritance stability is 2%), while the γ-plasmid has almost 100% stability. 相似文献
996.
997.
998.
An integrated flow injection process for analysis of intracellular components of microbes has been used to monitor plasmid content in Escherichia coli cultivations inoculated with cells subcultured in the presence or absence of ampicillin. The system allows sampling, sample handling, cell disruption, separation of intracellular components, and analysis in a semi-on-line mode of operation. The time scale for the assay is in the range 15 min (plasmid peak) to 25 min (complete assay cycle). As expected, lower initial plasmid content was found using an inoculum subcultured in the absence of ampicillin. More importantly, significant decrease in plasmid content was detected in the later stages of the cultivations (grown in ampicillin containing medium) even when using inoculum subcultured in the presence of ampicillin. This illustrates the versatility of the system, which allows monitoring of plasmid content as the cultivation proceeds. 相似文献
999.
Exocellular structures containing VirB2 proteins were, for the first time, localized on the surface of Agrobacterium by transmission electron microscopy. Using colloidal gold (CG)-labeled VirB2-specific antibodies, it was shown that VirB2 proteins enter into the composition of short surface pili, which emerge at the poles of acetosyringone (AS)-induced Agrobacterium cells. However, cells of the Ti plasmidless A. tumefaciens strain UBAPF-2 and cells not induced with AS were incapable of pilus synthesis. In suspension, mating Agrobacterium cells were connected together by short thick bridges. It was found that these bridges did not include as part of their structure CG-labeled VirB1 and VirB2 proteins. We did not find the tetracycline-resistant transconjugants after mating of A. tumefaciens donor cells harboring binary systems with plasmid-free A. tumefaciens GM-I 9023 in vir-induced and vir-uninduced conditions. However, the same strains can transfer pSUP106 plasmid via a vir-dependent way. We found that activated vir genes slightly stimulate pTd33 plasmid transfer via a tra-dependent pathway to plasmid-free strain UBAPF-2. It seems, that vir-induced T-DNA/plasmid DNA transfer machinery is not essential for the conjugation process between agrobacterial cells but may participate in this process. 相似文献
1000.