Transgenic Animals And Knockout Animals

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Transgenic Animals And Knockout Animals:

Transgenic Animals And Knockout Animals Dr. Amol Khanapure

Transgenic Animals:

Transgenic Animals A transgenic animal is one that carries a foreign gene that has been deliberately inserted into its genome. The foreign gene is constructed using Recombinant DNA Technology.

Different ways to create transgenic animals.:

Different ways to create transgenic animals. 1. Microinjection of recombinant DNA into the male pronucleus 2. embryonic stem cell transfer.

Transgene Construction:

Transgene Construction 3 parts of Transgene: Promoter sequence Transgene gene Termination sequence

Microinjection into the Male Pronucleus:

Microinjection into the Male Pronucleus 1. Most popular way 2. Eggs must be collected by super-ovulating female animals and these eggs must be fertilized in vitro.

Embryonic Stem Cell Manipulation:

Embryonic Stem Cell Manipulation This method is used mostly when trying to target a transgene to a specific site in the genome. Embryonic stem cells have the ability to differentiate into any of the three main germ layers. Once the transgene is incorporated into the ES cells, those cells can be left to divide in vitro, or they can be injected into a blastocyst and implanted into a host’s uterus to grow normally

PowerPoint Presentation:

4 approaches to use ES cells: Microinjection Introduction by virus Introduction by chemical Homologous recombination

Homologous Recombination.:

Homologous Recombination. The transgene is specifically targeted to a known location in the recipient genome, are introduced into the ES cells. Crossing over or homologous recombination can occur between the flanking DNA and the corresponding host chromosomal DNA, incorporating the transgene into a specific site dictated by the choice of flanking DNA. If these ES cells are then injected into blastocyst embryos in and implanted into the uterus of a foster mother they will grow normally.

Chemical And Viral Delivery:

Chemical And Viral Delivery Certain chemicals such as calcium phosphate or rubidium chloride and viruses will help carry the transgene into the cells. But similar to microinjection, the transgene will be placed randomly in the genome.

Screening For Transgenic Positives:

Screening For Transgenic Positives Screened by polymerase chain reaction (PCR) or by Southern blot analysis. Depending on the site of integration of the transgene, some transgenes may not be expressed if integrated into a transcriptionally inactive location. It is common practice to do further animal breeding to obtain maximal expression.

TRANSGENIC ANIMAL CLASSIFICATION AND EXAMPLES:

TRANSGENIC ANIMAL CLASSIFICATION AND EXAMPLES 5 Major Categories: Disease models Transpharmers Xenoplanters Food sources Scientific models

Disease Models:

Disease Models Disease models are animals that have been modified to exhibit the symptoms and progression of a particular disease, so that treatments for that disease can be tested on them . e.g. 1. AIDS Mouse 2. Alzheimer’s Mouse 3. Oncomouse 4. Parkinson’s Fly

Transpharmers:

Transpharmers Transpharmers are engineered to overexpress a particular gene in the mammary gland (so that the milk contains the desired protein). First done in an animal’s blood The trend has shifted to milk, since it is easier to acquire the drug from milk, and proteins expressed in milk are less likely to affect the animal’s physiology than in the blood. Promising, very expensive, low success rate Uses: Human lactoferrin, Human clotting factor

Xenotransplanters:

Xenotransplanters Xenotransplanters are being engineered to provide animal organs that are histocompatible with humans. The animal currently chosen for xenotransplant research is the pig since its physiology closely matches that of humans, and pigs are far cheaper than monkeys. Human trials have not yet been approved. Risk of Zoonotic infection.

Transgenic Food Sources:

Transgenic Food Sources Superpig Superfish

Transgenic Biological Models:

Transgenic Biological Models Made with the aim of increasing knowledge of genetics and expression, or some natural process. e.g. 1. ANDi 2. Smart Mouse 3. Supermouse 4. Youth Mouse 5. Influenza Resistant Mouse

PowerPoint Presentation:

The World's First Transgenic Monkey, ANDi. “Doogie” the transgenic smart mouse involved in a learning and memory test (Princeton, 2006).

Knockout Animals:

Knockout Animals A gene knockout is a genetic technique in which one of an organism's genes is made inoperative ("knocked out" of the organism). used in learning about a gene that has been sequenced, but which has an unknown or incompletely known function

Method To Create Knockout Animals:

Method To Create Knockout Animals A combination of techniques Beginning in the test tube with a plasmid, a bacterial artificial chromosome or other DNA construct, and proceeding to cell culture. Individual cells are genetically transfected with the DNA construct. Embryonic stem cells are genetically transformed and inserted into early embryos. Resulting animals with the genetic change in their germline cells can then often pass the gene knockout to future generations.

PowerPoint Presentation:

A laboratory mouse in which a gene affecting hair growth has been knocked out (left), is shown next to a normal lab mouse

PowerPoint Presentation:

A knockout mouse (left) that is a model of obesity, compared with a normal mouse

Uses:

Uses To understand the role of a specific gene or DNA region. Screening tools in the development of drugs. To target specific biological processes or deficiencies by using a specific knockout. T o understand the mechanism of action of a drug by using a library of knockout organisms spanning the entire genome, such as in Saccharomyces cerevisiae.

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