Transgenic mice.

Peter Ashby p-ashby at nimr.mrc.ac.uk
Wed Mar 12 12:42:50 EST 1997


In Article <5g6ekm$kqv at mserv1.dl.ac.uk>, "Joseph Cheer _ Coordinador de
Neurobiologia. FINC _" <cheer at javercol.javeriana.edu.co> wrote:

>Dear sirs,

There are a lot of people in the newsgroup who are not sirs.

>I am a Biologist who will start a PhD in Molecular Neuropharmacology 
>this fall. I am very interested in the use of transgenic mice (specially the 
>use of embryonic stem cells as a vehicle to mutate specific genes). I do 
>have some protocols but I have one question that seems to remain 
>unanswered. After pro nuclear microinjection how does the DNA template 
>integrate the host genome. Is it by homologous recombination? I would 
>like to know what the exact mechanism is. Could please shed some light in 
>this subject for me?

DNA introduced by pronuclear injection does not integrate by homologous
recombination otherwise a lot of sequences used would not integrate and
those that did would lead to knockouts.  If you could make knockouts with
pronuclear injection I would have a stack of Nature papers by now :-)

Transgenic constructs are generally inserted into one or more places in the
genome in a semi-random fashion.  They are in tandem head to tail arrays of
from one to dozens of copies, although one copy only can be very hard to
get.  I don't know for sure but I think they are usually blunted into place.

Disruptions caused by transgene insertions are usually because the transgene
has randomly inserted into coding sequence.

Hope this helps.

Peter


~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~
Peter Ashby                             National Institute for Medical Research
Eukaryotic Molecular Genetics           London, England
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