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Results for “vector engineering”

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At least 1,243 records · Page 69Linked to original sources
PubMed2004

Genetic engineering in the mouse: tuning TNF/TNFR expression.

With the exponential increase in the number of genes identified by various genome projects, it has become imperative that efficient methods be developed for deciphering gene function. Genetically engineered strains of mice are now critical research tools for basic biomedical research and for genomic approaches for the development of new therapeutic treatments for human disease. Over the past ten years it has become possible to make essentially any mutation in the mouse by transgenesis and homologous recombination in embryonic stem cells. Current advances in the "genetic engineering of the mouse," including the tissue-specific activation or inactivation of gene expression combined with developing technologies for switching gene expression on and off at will, provide experimental settings unprecedented in their potential to offer answers to long-standing questions.

Alleles↗
PubMed2005

Strategies for the use of site-specific recombinases in genome engineering.

Conventional gene targeting has been very useful in the study of gene function and regulation in mice. However, the methodologies involved have several limitations. First, mutations that cause embryonic lethality largely preclude studies of gene function at a later stage in development. Second, conditional and/or tissue-specific alterations of gene expression cannot be achieved using these methods. In addition, classical gene targeting can be difficult and time consuming. Strategies that make use of site-specific recombinases such as Cre and/or Flp have been developed in recent years to overcome these limitations. These new techniques include global and conditional knockouts, recombinase-mediated DNA insertion (RMDI), and recombinase-mediated cassette exchange (RMCE). Together, they have tremendously increased the number and variety of genetic manipulations that can be achieved.

Animals↗
PubMed2007

Fluorescent proteins and engineered cell lines.

Green fluorescent protein and other fluorescent proteins provide powerful tools for high content analysis of cellular processes. Engineering fluorescent protein sensors for expression in cellular assays requires consideration of a wide range of design factors to produce fusion proteins capable of generating informative and biologically relevant data while meeting the rigorous demands of high content screening. The target protein, fluorescent protein, host cell line, construct components and orientation, expression level, and other factors all contribute to the performance of the sensor. This chapter reviews the process of selecting an optimum combination of these design elements for engineering cells for high content analysis.

Animals↗
PubMed2006

Site-specific DNA excision in transgenic rice with a cell-permeable cre recombinase.

The removal of selected marker genes from transgenic plants is necessary to address biosafety concerns and to carry out further experiments with transgenic organisms. In the present study, the 12-amino-acid membrane translocation sequence (MTS) from the Kaposi fibroblast growth factor (FGF)-4 was used as a carrier to deliver enzymatically active Cre proteins into living plant cells, and to produce a site-specific DNA excision in transgenic rice plants. The process, which made cells permeable to Cre recombinase-mediated DNA recombination, circumvented the need to express Cre under spatiotemporal control and was proved to be a simple and efficient system to achieve marker-free transgenic plants. The ultimate aim of the present study is to develop commercial rice cultivars free from selected marker genes to hasten public acceptance of transgenic crops.

Blotting, Southern↗
PubMed1992

Construction of a plasmid containing the complete coding region of human elongation factor 2.

A plasmid pUChEF-2 containing the coding sequence as well as the complete 3'-untranslated region (3'UTR) of human EF-2 mRNA was constructed. The plasmid construct was assembled from a cDNA insert of pHGR81 (Rapp et al., (1988) Biol. Chem. Hoppe-Seyler 369, 247-250) comprising the C-terminal portion of the coding region and the 3'UTR, as well as a polymer chain reaction PCR fragment (Rapp et al., (1989) Biol. Chem. Hoppe-Seyler 370, 1071-1075) covering the missing part of the coding region from the amino-terminus.

Base Sequence↗
PubMed1992

Ecological studies on the intermediate host snails and the relevance to schistosomiasis control.

A detailed knowledge of distribution patterns of schistosome intermediate hosts and their population dynamics and factors affecting these patterns will provide useful information about the possibilities and desirability of conducting snail control measures in various transmission situations. On the basis of various case studies the association between the occurrence of human water contacts and the presence of schistosome intermediate hosts or infections in the intermediate hosts is illustrated. Other parameters affecting snail distribution patterns and density fluctuations are discussed. It is concluded that ecological studies on the intermediate hosts are extremely relevant, either to optimally apply existing control measures or to develop alternative measures of snail control, such as ecological or biological control.

Animals↗
PubMed2006

Baculovirus-mediated gene expression in chicken primary cells.

A recombinant baculovirus was constructed containing an expression cassette with a reporter gene, green fluorescent protein, directed by a constitutive mammalian promoter: a human cytomegalovirus immediate early promoter/enhancer (CMV-IE). High titer virus was prepared with ultracentrifugation. Efficient gene delivery and expression were observed in the virus-treated chicken primary culture, myoblast cells, and whole embryonic fibroblast cells. It was noticed that an addition of sodium butyrate (a selective histone deacetylase inhibitor) to viral transduction medium extremely enhanced the reporter-gene expression. However, there is no effect of presence of trichostatin A observed. To maximize the reporter-gene expression, the baculoviral infection condition was optimized with both cell types. Our approaches demonstrated that recombinant baculovirus could efficiently deliver its genome DNA into chicken primary cells and that CMV-IE, a mammalian-cell-active promoter, was functional in chicken primary cells and could direct a high level of gene expression. Clearly, the recombinant baculovirus provides an alternative means for foreign gene delivery into avian cells.

Animals↗