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Learn.genetics.utah --making a transgenic mouse



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.


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

so some one know about replacing silicon atom with phosphorous in semiconductors device?
s. Reply
how to fabricate graphene ink ?
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What is lattice structure?
s. Reply
of graphene you mean?
or in general
in general
Graphene has a hexagonal structure
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what is biological synthesis of nanoparticles
Sanket Reply
what's the easiest and fastest way to the synthesize AgNP?
Damian Reply
types of nano material
abeetha Reply
I start with an easy one. carbon nanotubes woven into a long filament like a string
many many of nanotubes
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I'm interested in nanotube
what is nanomaterials​ and their applications of sensors.
Ramkumar Reply
what is nano technology
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what is system testing?
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Victor Reply
Yes, Nanotechnology has a very fast field of applications and their is always something new to do with it...
Himanshu Reply
good afternoon madam
what is system testing
what is the application of nanotechnology?
In this morden time nanotechnology used in many field . 1-Electronics-manufacturad IC ,RAM,MRAM,solar panel etc 2-Helth and Medical-Nanomedicine,Drug Dilivery for cancer treatment etc 3- Atomobile -MEMS, Coating on car etc. and may other field for details you can check at Google
anybody can imagine what will be happen after 100 years from now in nano tech world
after 100 year this will be not nanotechnology maybe this technology name will be change . maybe aftet 100 year . we work on electron lable practically about its properties and behaviour by the different instruments
name doesn't matter , whatever it will be change... I'm taking about effect on circumstances of the microscopic world
how hard could it be to apply nanotechnology against viral infections such HIV or Ebola?
silver nanoparticles could handle the job?
not now but maybe in future only AgNP maybe any other nanomaterials
I'm interested in Nanotube
this technology will not going on for the long time , so I'm thinking about femtotechnology 10^-15
can nanotechnology change the direction of the face of the world
Prasenjit Reply
At high concentrations (>0.01 M), the relation between absorptivity coefficient and absorbance is no longer linear. This is due to the electrostatic interactions between the quantum dots in close proximity. If the concentration of the solution is high, another effect that is seen is the scattering of light from the large number of quantum dots. This assumption only works at low concentrations of the analyte. Presence of stray light.
Ali Reply
the Beer law works very well for dilute solutions but fails for very high concentrations. why?
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Smarajit Reply
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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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