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Inducible Gene Targeting in Mice Using the Cre/lox System.

Inducible Gene Targeting in Mice Using the Cre/lox System. - Protocols and Methods Forum

Inducible Gene Targeting in Mice Using the Cre/lox System. - Post Any Protocol, Method, Technique, Procedure or Tips / Troubleshooting for any Molecular Biology Technique.


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  #1  
Old 03-28-2007, 02:18 PM
S. Ottawa Grad Student
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Default Inducible Gene Targeting in Mice Using the Cre/lox System.




Hello Everyone,

I am new today to this group. I have looked at the previous postings I
could find regarding creating
KO mice using the Cre/Lox System and none of them seem to give me what
I need.

What I would like is a detailed, specific protocol for achieving this.
I have not been able to find such a document with sufficient technical
information regarding procedures, possible snags and useful tips on
troubleshooting techniques.

If anyone has such documents or knows where I can obtain them please
let me know.

Thank you to everyone in advance for any contributions.

S.

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  #2  
Old 03-29-2007, 09:35 AM
Kyle Legate
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Default Inducible Gene Targeting in Mice Using the Cre/lox System.

S. Ottawa Grad Student wrote:
For a good step-by-step guide to the embryonic stem cell work, you
should try to get a copy of "Integrin gene targeting" by Talts et al:

[Only registered users see links. ]

Additional information can be found in Manipulating the Mouse Embryo, by
Nagy et al. As for construction of the vector, I don't know of any
better way than to sit down with someone who has done it before. It's
easy to make a stupid mistake at this point that can cost months of time
(loxP sites in opposite orientations, for example). Perhaps if you join
the transgenic listserv and ask your question there someone will have
some literature to recommend.

What you can begin immediately, while you gather additional information
about how to construct the vector, is to screen a commercial genomic
library using a fragment of your gene of interest as a Southern probe.
The result will tell you which PAC or BAC clone to obtain from the
company to use as starting material to make your targeting construct.
You will need around 20 kb for both recombination arms, and they must
have no mutations (lest you lose most of your targeting efficiency) so I
wouldn't even consider trying to obtain it through in-house PCR.
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Old 01-21-2010, 02:23 AM
Pipette Filler
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Default Re: Inducible Gene Targeting in Mice Using the Cre/lox System.

hello,

I have been looking for mutant loxp sites online for a few days, so far I know that loxp 2272, loxp 171, loxp 511, loxp 512 are all mutant types of loxp; does anyone know about the other mutant loxp sites? I am appreciated for letting me know about it. thank you

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cre or lox , gene , inducible , mice , system , targeting


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