transgenic animals

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history : 

history Studying Earliest history among farmers-selective breeding Discovery of DNA-1953 New molecular biology techniques-Rdna,genomic mapping, genetic cloning


TRANSGENESIS Radical new technology-direct manipulation of genetic material Term transgenic-J W Gordon and F H Rudell “Tailor-made animals”-wanted characteristics More efficient than selective breeding Cross species More economical-fewer feed supplements /medical treatments

Slide 4: 

A transgenic animal is one that carries a foreign gene that has been deliberately inserted into its genome. Genetically modified organism GMO The foreign gene is constructed using recombinant DNA technology First transgenic animal produced = Founder Animal

Transgenic structure : 

Transgenic structure Promoter Coding sequence Poly A tail

Transgenic animals : 

Transgenic animals Mice rabbit Sheep Goats Cow Pigs Chicken Fish fly

Timeline… : 


Growth hormone gene transfer : 

Growth hormone gene transfer 1982-Brinster and Palmiter successfully injected into the eggs of a mouse the gene which controls the expression of the growth hormone

Mice as ideal transgenic animals : 

Mice as ideal transgenic animals The first successful transgenic animal was a mouse 95% of transgenic animals used in biomedical research 80% of mouse genes function the same as those in humans. Mice also have a short reproduction cycle and their embryos are amenable to manipulation.

Slide 10: 

To date, there are three basic methods of producing transgenic animals: DNA microinjection Retrovirus-mediated gene transfer Embryonic stem cell-mediated gene transfer

DNA microinjection : 

DNA microinjection predominant method used to produce transgenic farm animals. Since the insertion of DNA results in a random process, transgenic animals are mated to ensure that their offspring acquire the desired transgene Success rates 10-30%

Establishing transgenic mice with DNA microinjection : 

Establishing transgenic mice with DNA microinjection

Analysis of DNA integration : 

Analysis of DNA integration 3-4 wk old mice Piece of tail-DNA testing Detection of transgene by PCR

Slide 16: 

Advantages: No theoretical size of the construct Can be applied to other species Disadvantages: Genes can only be added not deleted Small percentage are transgenic and only a proportion express the gene in high levels

Retrovirus mediated gene transfer : 

Retrovirus mediated gene transfer retroviruses used as vectors to transfer genetic material into the host cell, resulting in a chimera, an organism consisting of tissues or parts of diverse genetic constitution chimeras are inbred for as many as 20 generations until homozygous (carrying the desired transgene in every cell) transgenic offspring are born Lentivirus mediated gene transfer-latest advancement

advantages : 

advantages Earliest method of gene transfer in mammals Killed virus-replication defect Transfection property Wide range of cells can be transfected Successful method-chicken,fish,monkey Transgene size-<8kb

Chimeric mouse : 

Chimeric mouse

Embryonic Stem Cell-Mediated Gene Transfer : 

Embryonic Stem Cell-Mediated Gene Transfer isolation of totipotent stem cells (stem cells that can develop into any type of specialized cell) from embryos the desired gene is inserted into these cells cells containing the desired DNA are incorporated into the host’s embryo, resulting in a chimeric animal Allows testing for transgenes at cell stage Less random insertion than the other two methods

Transgenic animals-Engineered embyronic stem cell method (used for gene knockouts) : 

Transgenic animals-Engineered embyronic stem cell method (used for gene knockouts)

Littermates : 

Littermates Black mouse - no apparent ES cell contribution Chimeric founder - strong ES cell contribution Chimeric founder - weaker ES cell contribution

Embryonic stem cells : 

Embryonic stem cells 1.May be transfected with transgene 2.Genes can be removed(knockout) 3.Gene targetting(homologous recombination)

Gene knockout : 

Gene knockout

Slide 27: 

The first knockout mouse was created by Mario R. Capecchi, Martin Evans and Oliver Smithies in 1989, for which they were awarded the Nobel Prize for Medicine in 2007 the p53 knockout mouse is named after the p53 gene which codes for a protein that normally suppresses the growth of tumors by arresting cell division.

Slide 28: 

A humouse is an immunodeficient mouse reconstituted with a human immune system Nude mouse-genetically engineered mouse with absent thymus

Nuclear transfer : 

Nuclear transfer Somatic cells are transfected or genetically altered prior to nuclear transfer DOLLY-1997 100% certainty of transgene in progeny Used in production of transgenic livestock

Slide 32: 

Denucleus oocyte by mechanical removal Recepient oocyte for embryo cloning

Other methods : 

Other methods Sperm mediated gene transfer-sperm co incubated with gene construct and introduced using AI/IVF methods Artificial chromosome mediated transfer: Yeast artificial chromosome(YAC) used to hold large DNA fragments

Transgenic rabbits : 

Transgenic rabbits Human Alfa antitrypsin,IL-2,tpa,superoxide dismutase Human disease model-atherosclerosis Replication of HIV Disadvantages- Sexual maturation-months Large size costly

Transgenic sheep and goat : 

Transgenic sheep and goat Antithrombin III(A Tryn-2006) T-pa Mab Alfa 1 antitrypsin

Famous sheep : 

Famous sheep 1990-tracey -human alfa antitrypsin 1995-megan and morag –cloned 1996-dolly-cloned 1997-polly-first cloned animal containing a human gene expressing factor 9

Transgenic chickens : 

Transgenic chickens Used to synthesize several important human proteins in the white of their eggs Mab,lyzozyme,GH,insulin

Transgenic pigs : 

Transgenic pigs Improved productivity-GH Increase disease resistance Human disease models-retinitis pigmentosa Biomedical products-human haemoglobin,protein C,Xenotransplantation

“Enviropigs” : 

“Enviropigs” Transgenic pigs expressing the phytase gene in their salivary glands The phytase gene was introduced via DNA microinjection and used the parotid secretory protein promoter to specifically drive expression in the salivary glands Phytate is the predominant storage form of phosphorus in plant-based animal feeds (e.g., soybean meal) Pigs and poultry cannot digest phytate and consequently excrete large amounts of phosphorus “Enviro-pigs” excrete 75% less phosphorus EnviropigTM an environmentally friendly breed of pigs that utilizes plant phosphorus efficiently.

Transgenic fish : 

Transgenic fish The growth hormone is expressed by a metallothionein promoter. The hormone gene and the promoter (from the metallothionein gene) are cloned from the sockeye salmon. Result- Transgenic fish are 10-30 times larger than normal fish

Slide 41: 

Drosophila melanogaster- Short life span Low maintainance Simple genome Ideal model to study the Effect of genetic changes on development

Use of transgenic animals : 

Use of transgenic animals Agriculture-breeding,quality,disease resistant

Slide 43: 

Medicine- Proteins of human importance Disease models-oncomouse Human gene therapy xenotransplantation

Transgenic mice models for human disease : 

Transgenic mice models for human disease

Another Transgenic mouse application: Marathon Mice : 

Another Transgenic mouse application: Marathon Mice Instead of improving times by fractions of a second, the genetically enhanced “marathon” mice (above, on the treadmill in San Diego) ran twice as far and nearly twice as long as ordinary rodents. The peroxisome proliferator-activated receptor (PPAR-delta) gene was overexpressed in these transgenic mice. For details, see Dr. Ron Evans and one of his genetically engineered “marathon” mice. The enhanced PPAR-delta activity not only increased fat burning, but transformed skeletal muscle fibers, boosting so-called "slow-twitch" muscle fibers, which are fatigue resistant, and reducing 'fast-twitch' fibers, which generate rapid, powerful contractions but fatigue easily.

Human proteins from animals(pharming) : 

Human proteins from animals(pharming)

first transgenic mammal : 

first transgenic mammal Herman the Bull (Lelystad, 16 December 1990 – Leiden, 2 April 2004) was the first genetically modified or transgenic mammal in the world Scientists microinjected cells with the human gene coding for lactoferrin

Slide 48: 

In 1997,the first transgenic cow Rosie produced human protein enriched milk at 2.4gm/litre Contains the human gene alfa lactalbumin Other transgenic cows produce milk with Less lactose or cholesterol

industrial : 

industrial In 2001, two scientists at Nexia Biotechnologies in Canada spliced spider genes into the cells of lactating goats. The goats began to manufacture silk along with their milk and secrete tiny silk strands from their body by the bucketful.

problems : 

problems Abnormalities suffered are more Reduced fertility Oversize foetus Respiratory/circulatory problems Weak immune systems Age faster-short telomere Those that do not express the gene-destroyed

Religious ethical considerations : 

Religious ethical considerations Man "playing the role of God" Manipulating nature Consumer's safety  Boycott on transgenic products unnatural and unsafe

Environmental issues : 

Environmental issues Changing the food/web chain Depopulation amongst native species Increase of population amongst transgenic animals Severe changes within the environments such as depopulation of certain species of plant, increase in allergies, mixing of natural and transgenic animals, etc. Changing the direction of evolution?

And now there is pet cloning for a “small” fee… : 

And now there is pet cloning for a “small” fee… Nine-week-old "Little Nicky" peers out from her carrying case in Texas. Little Nicky, a  cloned cat, was sold to its new owner by Genetic Savings and Clone for $50,000 in December 2004. August 07, 2008 | Bernann McKinney with one of the 5 puppies cloned from Booger, her late pet pit bull. It cost her $50,000. When Booger was diagnosed with cancer, a grief-stricken McKinney sought to have him cloned -- first by the now-defunct Genetic Savings and Clone, and then by South Korean company RNL Bio.

And then there is “transgenic art” with GFP… : 

And then there is “transgenic art” with GFP…

Transgenic plants : 

Transgenic plants possess several desirable traits resistance to pests, herbicides harsh environmental conditions improved product shelflife increased nutritional value Bt-maize –spliced toxin producing gene from bacteria-sicken insects that try to consume it

Golden rice : 

Golden rice

Human clones…. : 

Human clones….

Pros and cons : 

Pros and cons Biologically related children(infertile couples) Avoid genetic disease Rejection proof transplants Replicate a loved one Medical research Rights of a clone? Safety aspect? Humans as commodities



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