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Delivery of Genes to the Eye Using Lentiviral Vectors

The primary aim of gene transfer into the retinal cells has been to investigate the developmental mechanisms of the retinal cells or to reverse retinal diseases. Retroviruses have been used to investigate the differentiation of retinal cells, to study the embryonic retina in vivo or explant o ...

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Lentiviral Transduction of Human Dendritic Cells

Dendritic cells (DCs) are potent antigen-presenting cells (APCs) that play a pivotal role in stimulating antigen-specific T cells in vivo (1,2). The cardinal properties of DCs are: (1) the ability to take up, process, and present antigens; (2) the ability to migrate through different tissues into ...

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Gene Transfer by Retroviral Vectors: An Overview

Viruses have evolved to deliver their genetic cargo to cells and, due to the pathogenicity of some viruses, this process has been the subject of a great deal of study. In this respect, retroviruses came to the fore in the early 1900s with the demonstration by Ellermann and Bang (1908) and by Rous (1911) that chi ...

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Gene Delivery to Cells in Culture Using Retroviruses

Moloney leukemia virus-based vectors can be generated in cells that express the products of three retroviral genes, gag, pol, and env. There are a number of cell lines such as PG13 (1) and FLYA13 (2), known as packaging cells, that have been established that stably express these genes. When these cells are t ...

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Retrovirus-Mediated Gene Transfer to Tumors: Utilizing the Replicative Power of Viruses to Achieve Highly Efficient Tumor Transduction In Vivo

Vectors derived from retroviruses have been widely studied as tools for gene transfer into mammalian tissue in vivo. One application for which retroviral vectors have received particular attention is gene transfer into tumor cells for treatment of cancer. Simple retroviruses, such as ...

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Dilivery of Genes to Hematopoietic Stem Cells

Bone marrow hematopoiesis is maintained by hematopoietic stem cells (HSC) (1). Because of their unique features to self-renew and differentiate along all lineages of hematopoietic cells, even a single HSC can completely reconstitute bone marrow hematopoiesis of irradiated recip ...

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Systemic First-Line Phenotyping

 With the completion of the mouse genome sequence an essential task for biomedical sciences in the twenty-first century will be the generation and functional analysis of mouse models for every gene in the mammalian genome. More than 30,000 mutations in ES cells will be engineered and thousands of ...

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Pathologic Phenotyping of Mutant Mice

The easiest and cheapest way to analyze the phenotype of most knockout mice is to do a comprehensive necropsy and histopathologic examination of slides of all tissues. Once any lesion is found in a knockout mouse a vast contemporary and traditional literature can be searched for occurrences of s ...

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Influence of Genetic Background on Genetically Engineered Mouse Phenotypes

The history of mouse genetics, which involves the study of strain-dependent phenotype variability, makes it clear that the genetic background onto which a gene-targeted allele is placed can cause considerable variation in genetically engineered mouse (GEM) phenotype. This variat ...

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Sperm Cryopreservation and In Vitro Fertilization

Since the mouse has become the most profound model system to investigate the genetics and pathogenetics of human diseases, a huge number of new mutant mouse strains has been generated and still a lot effort is being done to increase the number of suitable mouse models. In nearly all animal facilities t ...

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Lentiviral Transgenesis

Lentiviral vectors efficiently transfer genes into a broad spectrum of cells and tissues, including non-dividing cells and stem cells. Lentiviruses integrate their viral genome into the host chromosome, which is the basis for virus latency as well as stable transgene expression. A rath ...

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Transposon Mutagenesis in Mice

Understanding the functional landscape of the mammalian genome is the next big challenge of biomedical research. The completion of the first phases of the mouse and human genome projects, and expression analyses using microarray hybridization, generate critically important que ...

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Inducible Cre Mice

The Cre/lox site-specific recombination system has emerged as an important tool for the generation of conditional somatic mouse mutants. This method allows one to control gene activity in space and time in almost any tissue of the mouse, thus opening new avenues for studying gene function and for ...

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Generation of Cre Recombinase-Expressing Transgenic Mice Using Bacterial Artificial Chromosomes

Generation of genetically modified mice is one of the primary methods for understanding gene function. In particular, approaches that allow for restricting the effects of a mutation to defined cell-types are fundamental for understanding the roles of genes in specific cells or tissues. T ...

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VelociMouse: Fully ES Cell-Derived F0-Generation Mice Obtained from the Injection of ES Cells into Eight-Cell-Stage Embryos

With the completion of the human and mouse genome sequences and the development of high-throughput knockout mouse technologies, there is now a need for equally high-throughput methods for the production of mice for phenotypic studies. In response to this challenge, we recently developed a ...

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Aggregation Chimeras: Combining ES Cells, Diploid, and Tetraploid Embryos

During the past 40 years, mouse chimeras have served as invaluable tools for studying not only genetics but also embryonic development, and the path from undifferentiated cell populations to fully committed functional cell types. This chapter gives a description of the early events of cell ...

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Isolation, Microinjection and Transfer of Mouse Blastocysts

Genetically modified mice by means of homologous recombination in embryonic stem (ES) cells are generated by injection of manipulated ES cells into recipient blastocysts. The injected blastocysts, following reintroduction into recipient foster mice, will produce chimeric mi ...

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Cloning of ES Cells and Mice by Nuclear Transfer

We have been able to develop a stable nuclear transfer (NT) method in the mouse, in which donor nuclei are directly injected into the oocyte using a piezo-actuated micromanipulator. Although the piezo unit is a complex tool, once mastered it is of great help not only in NT experiments, but also in almost all o ...

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Differentiation Analysis of Pluripotent Mouse Embryonic Stem (ES) Cells In Vitro

Pluripotent embryonic stem (ES) cells are characterized by their almost unlimited potential to self-renew and to differentiate into virtually any cell type of the organism. Here we describe basic protocols for the in vitro differentiation of mouse ES cells into cells of the cardiac, neuron ...

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ES Cell Line Establishment

A method is described to establish mouse embryonic stem cell (ESC) lines from hybrid and inbred strains of mice. Attention is paid not only to the methodology for isolation and culture but also to the validation of freshly derived lines, in order to be maintained for prolonged time without signific ...

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