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Chromosomes, Artificial, Bacterial: DNA constructs that are composed of, at least, a Replication origin, for successful replication, propagation to and maintenance as an extra chromosome in bacteria. In addition, they can carry large amounts (about 200 kilobases) of other sequence for a variety of bioengineering purposes.
 JoVE Neuroscience

Detection of the Genome and Transcripts of a Persistent DNA Virus in Neuronal Tissues by Fluorescent In situ Hybridization Combined with Immunostaining

1Virus and Centromere Team, Centre de Génétique et Physiologie Moléculaire et Cellulaire, CNRS UMR 5534, 2Université de Lyon 1, 3Laboratoire d'excellence, LabEX DEVweCAN, 4Institut de Virologie Moléculaire et Structurale, CNRS UPR 3296, 5Centre de Recherche en Cancérologie de Lyon, INSERM U1052, CNRS UMR 5286


JoVE 51091

 JoVE Developmental Biology

In Vitro Colony Assays for Characterizing Tri-potent Progenitor Cells Isolated from the Adult Murine Pancreas

1Diabetes and Metabolism Research Institute, Beckman Research Institute of City of Hope, 2Irell & Manella Graduate School of Biological Sciences, Beckman Research Institute of City of Hope, 3Division of Chemistry and Chemical Engineering, California Institute of Technology


JoVE 54016

 Science Education: Essentials of Genetics

Recombineering and Gene Targeting

JoVE Science Education

One of the most widely used tools in modern biology is molecular cloning with restriction enzymes, which create compatible ends between DNA fragments that allow them to be joined together. However, this technique has certain restrictions that limit its applicability for large or complex DNA construct generation. A newer technique that addresses some of these shortcomings is recombineering, which modifies DNA using homologous recombination (HR), the exchange between different DNA molecules based on stretches of similar or identical sequences. Together with gene targeting, which takes advantage of endogenous HR to alter an organism’s genome at a specific loci, HR-based cloning techniques have greatly improved the speed and efficacy of high-throughput genetic engineering.In this video, we introduce the principles of HR, as well as the basic components required to perform a recombineering experiment, including recombination-competent organisms and genomic libraries such as bacterial artificial chromosomes (BAC). We then walk through a protocol that uses recombineering to generate a gene-targeting vector that can ultimately be transfected into embryonic stem cells to generate a transgenic animal. Finally, several applications that highlight the utility and variety of recombineering techniques wi

 JoVE Medicine

Transposon Mediated Integration of Plasmid DNA into the Subventricular Zone of Neonatal Mice to Generate Novel Models of Glioblastoma

1Department of Neurosurgery, University of Michigan School of Medicine, 2Department of Pediatrics, Division of Hematology-Oncology, University of Michigan School of Medicine, 3Department of Cell and Developmental Biology, University of Michigan


JoVE 52443

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 JoVE Biology

Quantitation and Analysis of the Formation of HO-Endonuclease Stimulated Chromosomal Translocations by Single-Strand Annealing in Saccharomyces cerevisiae

1Irell & Manella Graduate School of Biological Sciences, 2Department of Molecular and Cellular Biology, City of Hope Comprehensive Cancer Center and Beckman Research Institute, 3Department of Biochemistry and Molecular Biology, University of Southern California, Norris Comprehensive Cancer Center


JoVE 3150

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 JoVE Immunology and Infection

Phage Phenomics: Physiological Approaches to Characterize Novel Viral Proteins

1Department of Biology, San Diego State University, 2Computational Science Research Center, San Diego State University, 3Bioinformatics and Medical Informatics Research Center, San Diego State University, 4Department of Mathematics and Statistics, San Diego State University, 5Department of Computer Science, San Diego State University, 6Mathematics and Computer Science Division, Argonne National Laboratory, 7SPARC Committee, Broad Institute


JoVE 52854

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 JoVE Medicine

Cell Death Associated with Abnormal Mitosis Observed by Confocal Imaging in Live Cancer Cells

1Cancer Research Center, Sheba Medical Center, 2The Neufeld Cardiac Research Institute, Tel-Aviv University, 3Department of Physiology and Pharmacology, Tel-Aviv University, 4Imaging Unit, Sackler Faculty of Medicine, Tel-Aviv University, 5Biotechnology and Cell Signaling, Ecole Superieure de Biotechnologie Strasbourg, 6Department of Human Molecular Genetics and Biochemistry, Tel-Aviv University


JoVE 50568

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