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Learn.genetics.utah --making a transgenic mouse

Source

http://learn.genetics.utah.edu/content/science/transgenic/

Mario caoechhi

Mario R. Capecchi, Ph.D., of the University of Utah, won the 2007 Nobel Prize in Physiology or Medicine. Capecchi shared the prize with Oliver Smithies of University of North Carolina, Chapel Hill, and Sir Martin Evans of Cardiff University in the UK.

The prize recognized Capecchi's pioneering work on "knockout mouse" technology, a gene-targeting technique that has revolutionized genetic and biomedical research, allowing scientists to create animal models for hundreds of human diseases.

As a child, Capecchi wandered homeless in Italy. As a researcher, his first attempts at gene targeting were deemed not ready for funding by the National Institutes of Health. Capecchi is an individual whose personal life proves that while some events are not probable, anything is possible.  Read Mario's story .

During the 1980s, Capecchi devised a way to change or remove any single gene in the mouse genome, creating strains of mice that pass the altered gene from parent to offspring. In the years since, these "transgenic" and "knockout" mice have become commonplace in the laboratory.

Capecchi's pioneering work in gene targeting has taught us much about how the body builds—and rebuilds—itself. He has given scientists worldwide the tools to make important discoveries about human diseases, from cancer to obesity.

And he has raised a key question for the future of human medicine: if we can replace a perfectly good gene with a mutated one, can we also go the other way, replacing problem genes with those that work?

Mario’s lab at work

What makes an arm an arm? Capecchi's research team is working on answering that question using gene targeting. They have systematically "knocked out" a set of genes in mice, called homeotic genes, which govern body patterning during development. For example, one of the lab's most recent genetic discoveries may explain  why we lack spare ribs . Find out more about how homeotic genes work in Genes Determine Body Patterns .

YOUR GOAL: You are studying how a particular gene, named OhNo, might play a role in panic attacks. You want to study mice that are missing this gene. To "knockout" the OhNo gene, you will replace it with a mutated copy that doesn't work.

Here's how:

1. Isolate Stem Cells

Isolate embryonic stem cells that originated from male brown mice with a normal OhNo gene (blue).

2. Add Inactive Gene With Marker

To these cells, add a copy containing a mutated, inactive OhNo gene (red), and a drug resistance marker gene (p3. Similar Genes Naturally Swap

3. Similar Genes Naturally Swap

By mechanisms that are not completely understood yet, similar genes will swap places. The OhNo gene plus drug resistance marker gene is incorporated into the genome, and the normal version is kicked out. This process is called homologous recombination.

ink).

4. Add Drug

Cells that haven't incorporated the inactive OhNo gene don't have the drug resistance marker gene (pink).

Adding the drug kills cells without the marker, leaving you with only cells that have an inactive version of the OhNo gene.

5. Grow Chimeric Mice

By transplanting stem cells that carry the inactive ohNo gene into a white mouse embyro, you'll create what is called a chimera. Chimeras have patches of cells throughout their bodies that grew from white mouse cells and patches that grew from brown stem cells. Some of the cells that have the inactive OhNo gene may develop into reproductive cells.

Chimeras are easy to identify because they have both brown and white patches of fur.

6. Mate Male Chimera

If a male chimera has some reproductive cells (sperm) that originated from the brown stem cells, he will produce some brown offspring when mated with a white female.

7. Test and Breed Brown Offspring

Half of the brown offspring will have a copy of the inactive OhNo gene in all of their cells—including their reproductive cells. These mice have one normal copy of the OhNo gene from their

mother (not shown) and one inactive copy from their father. So half of their reproductive cells will contain a normal copy, and half will contain an inactive copy.

These mice can be identified by performing DNA sequencing in their OhNo genes and then bred with each other.

8. You've Made a Knockout Mouse

One fourth of your resulting offsping will have two copies of the "knocked-out" or inactive OhNo gene. You can now study these mice to determine how lacking the OhNo gene may affect panic attacks.

Supported by a Science Education Partnership Award (SEPA) Grant No. R25RR023288 from the National Center for Research Resources, a component of the NIH. The contents provided here are solely the responsibility of the authors and do not necessarily represent the official views of NIH.

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Questions & Answers

Is there any normative that regulates the use of silver nanoparticles?
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Damian Reply
research.net
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Introduction about quantum dots in nanotechnology
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Devang Reply
are you nano engineer ?
s.
fullerene is a bucky ball aka Carbon 60 molecule. It was name by the architect Fuller. He design the geodesic dome. it resembles a soccer ball.
Tarell
what is the actual application of fullerenes nowadays?
Damian
That is a great question Damian. best way to answer that question is to Google it. there are hundreds of applications for buck minister fullerenes, from medical to aerospace. you can also find plenty of research papers that will give you great detail on the potential applications of fullerenes.
Tarell
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Abhijith Reply
Mostly, they use nano carbon for electronics and for materials to be strengthened.
Virgil
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CYNTHIA
carbon nanotubes has various application in fuel cells membrane, current research on cancer drug,and in electronics MEMS and NEMS etc
NANO
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s. Reply
Yeah, it is a pain to say the least. You basically have to heat the substarte up to around 1000 degrees celcius then pass phosphene gas over top of it, which is explosive and toxic by the way, under very low pressure.
Harper
Do you know which machine is used to that process?
s.
how to fabricate graphene ink ?
SUYASH Reply
for screen printed electrodes ?
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What is lattice structure?
s. Reply
of graphene you mean?
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or in general
Ebrahim
in general
s.
Graphene has a hexagonal structure
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Source:  OpenStax, Open genetics. OpenStax CNX. Jan 08, 2015 Download for free at https://legacy.cnx.org/content/col11744/1.3
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