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Genetic Engineering: Directed modification of the gene complement of a living organism by such techniques as altering the DNA, substituting genetic material by means of a virus, transplanting whole nuclei, transplanting cell hybrids, etc.

Genetic Engineering of an Unconventional Yeast for Renewable Biofuel and Biochemical Production

1Department of Biochemistry, Yong Loo Lin School of Medicine, National University of Singapore, 2NUS Synthetic Biology for Clinical and Technological Innovation (SynCTI), Life Sciences Institute, National University of Singapore, 3Food Science and Chemical Engineering, Singapore Institute of Technology

JoVE 54371


 Genetics

An Overview of Genetic Engineering

JoVE 5552

Genetic engineering – the process of purposefully altering an organism’s DNA – has been used to create powerful research tools and model organisms, and has also seen many agricultural applications. However, in order to engineer traits to tackle complex agricultural problems such as stress tolerance, or to realize the promise of gene therapy for treating human diseases, further advances in the field are still needed. Important considerations include the safe and efficient delivery of genetic constructs into cells or organisms, and the establishment of the desired modification in an organism’s genome with the least “off-target” effects. JoVE’s Overview of Genetic Engineering will present a history of the field, highlighting the discoveries that confirmed DNA as the genetic material and led to the development of tools to modify DNA. Key questions that must be answered in order to improve the process of genetic engineering will then be introduced, along with various tools used by genetic engineers. Finally, we will survey several applications demonstrating the types of experimental questions and strategies in the field today.


 Genetics

Genetic Engineering of Model Organisms

JoVE 5327

Transgenesis, or the use of genetic engineering to alter gene expression, is widely used in the field of developmental biology. Scientists use a number of approaches to alter the function of genes to understand their roles in developmental processes. This includes replacement of a gene with a nonfunctional copy, or adding a visualizable tag to a gene that allows the resultant fusion protein to be tracked throughout development. In this video, the viewers will learn about the principles behind transgenesis, as well as the basic steps for introducing genetic constructs into an animal and targeting genes of interest. This is followed by the discussion of a protocol to create knockout mice. Lastly, some specific applications of transgenic technologies in the field of developmental biology will be reviewed.


 Developmental Biology

Genetic Manipulation of the Plant Pathogen Ustilago maydis to Study Fungal Biology and Plant Microbe Interactions

1Institute for Microbiology, Heinrich-Heine University Düsseldorf, 2Bioeconomy Science Center (BioSC), 3Department of Genetics, Institute of Applied Biosciences, Karlsruhe Institute of Technology, 4Cluster of Excellence in Plant Sciences (CEPLAS), Heinrich-Heine University Düsseldorf

JoVE 54522


 Genetics

Preparation and In Vitro Characterization of Magnetized miR-modified Endothelial Cells

1Reference and Translation Center for Cardiac Stem Cell Therapy (RTC), Department of Cardiac Surgery, University of Rostock, 2Physikalisch-Technische Bundesanstalt, 3Department of Radiology and Neuroradiology, Ernst-Moritz-Arndt-University Greifswald, 4Electron Microscopy Center, University of Rostock

JoVE 55567


 Medicine

Protocol for MicroRNA Transfer into Adult Bone Marrow-derived Hematopoietic Stem Cells to Enable Cell Engineering Combined with Magnetic Targeting

1Reference and Translation Center for Cardiac Stem Cell Therapy (RTC), Department of Cardiac Surgery, Rostock University Medical Center, 2Department Life, Light and Matter of the Interdisciplinary Faculty, Rostock University

Video Coming Soon

JoVE 57474


 JoVE In-Press

Efficient Generation of hiPSC Neural Lineage Specific Knockin Reporters Using the CRISPR/Cas9 and Cas9 Double Nickase System

1Department of Neurosurgery, The University of Texas Health Science Center at Houston, 2Center for Stem Cell and Regenerative Medicine, Brown Foundation Institute of Molecular Medicine, The University of Texas Health Science Center at Houston, 3The Senator Lloyd & B. A. Bentsen Center for Stroke Research, Brown Foundation Institute of Molecular Medicine, The University of Texas Health Science Center at Houston, 4Summer Research Program, Office of Educational Programs, The University of Texas Health Science Center at Houston, 5Department of Anesthesiology, Shengjing Hospital, China Medical University, 6Department of Oncology, Renji Hospital, Shanghai Jiaotong University School of Medicine, 7Biology Department, University of West Georgia

JoVE 52539


 Developmental Biology

The Arteriovenous (AV) Loop in a Small Animal Model to Study Angiogenesis and Vascularized Tissue Engineering

1Department of Plastic and Hand Surgery and Laboratory for Tissue Engineering and Regenerative Medicine, University Hospital of Erlangen, Friedrich-Alexander University of Erlangen-Nürnberg (FAU), 2Genetic Engineering and Biotechnology Institute for Postgraduate Studies, Baghdad University, 3Department of Plastic, Hand and Microsurgery, Sana Klinikum Hof GmbH

JoVE 54676


 Bioengineering

CRISPR/Cas9-Mediated Targeted Integration In Vivo Using a Homology-Mediated End Joining-Based Strategy

1Institute of Neuroscience, State Key Laboratory of Neuroscience, Key Laboratory of Primate Neurobiology, CAS Center for Excellence in Brain Science and Intelligence Technology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, 2College of Life Sciences, University of Chinese Academy of Sciences, 3School of Life Science and Technology, Shanghai Tech University, 4Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences

Video Coming Soon

JoVE 56844


 JoVE In-Press

Evaluation of Vascular Control Mechanisms Utilizing Video Microscopy of Isolated Resistance Arteries of Rats

1Department of Physical Therapy, Marquette University, 2Medical College of Wisconsin, 3Department of Physiology, Medical College of Wisconsin, 4Graduate Programs of Nurse Anesthesia, Texas Wesleyan University, 5Office of Research, Medical College of Wisconsin

JoVE 56133


 Medicine

Lignin Down-regulation of Zea mays via dsRNAi and Klason Lignin Analysis

1The School of Plant Sciences, University of Arizona, 2Department of Chemical Engineering and Materials Science, DOE Great Lakes Bioenergy Research Center, Michigan State University, 3The Institute for Sustainable and Renewable Resources, The Institute for Advanced Learning and Research, 4Department of Plant, Soil and Microbial Sciences, Michigan State University

JoVE 51340


 Bioengineering

Recombineering and Gene Targeting

JoVE 5553

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


 Genetics

Clinical Application of Sleeping Beauty and Artificial Antigen Presenting Cells to Genetically Modify T Cells from Peripheral and Umbilical Cord Blood

1Division of Pediatrics, U.T. MD Anderson Cancer Center, 2Department of Stem Cell Transplantation and Cellular Therapy, U.T. MD Anderson Cancer Center

JoVE 50070


 Immunology and Infection

Detecting Estrogenic Ligands in Personal Care Products using a Yeast Estrogen Screen Optimized for the Undergraduate Teaching Laboratory

1Department of Biology, University of the South, 2School of Biological Sciences, Louisiana Tech University, 3School of Medicine, Louisiana State University Health Sciences Center, 4Department of Biology, Furman University, 5Department of Computer Science, Louisiana Tech University, 6Clemson University

JoVE 55754


 Biology

In Vitro Growth of Mouse Preantral Follicles Under Simulated Microgravity

1Reproductive Medicine Center, The First Affiliated Hospital of Wenzhou Medical University, 2Department of Orthopaedics, The Second Affiliated Hospital of Wenzhou Medical University, 3Department of Obstetrics, The First Affiliated Hospital of Wenzhou Medical University, 4School of Laboratory Medicine and Life Science, Wenzhou Medical University, 5School of Pharmaceutical Science, Wenzhou Medical University, 6Stem Cells and Genetic Engineering Group, AgriBioscience Research Centre, Department of Economic Development, Jobs, Transport and Resources, 7Department of Histology and Embryology, Wenzhou Medical University

JoVE 55641


 Developmental Biology

Fundamentals of Breeding and Weaning

JoVE 10293

Source: Kay Stewart, RVT, RLATG, CMAR; Valerie A. Schroeder, RVT, RLATG. University of Notre Dame, IN

Millions of mice and rats are bred for use in biomedical research each year. Worldwide, there are several large commercial breeding facilities that supply mice to research laboratories, but many facilities choose to also breed mice and rats in-house to reduce costs and increase research options. When breeding in the animal facility, researchers are able to manipulate the genetics of the animals, time the pregnancies to meet the needs of the research, and work with embryos and neonates as required. Mice and rats can be bred in a variety of schemes and methods. Technical procedures, such as the use of vaginal cytology, visualization of the vaginal area, and observation of copulatory plugs, have been developed to assist with the synchronization of breeding to correspond to research requirements. This manuscript is an overview of the basic fundamentals of mouse and rat breeding and technical procedures used. More detailed descriptions of the complex breeding schemes, and the full description of the methods for vaginal cytology, are available in the list of references.


 Lab Animal Research

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