In order to make a directed genetic change in a specific mouse gene we exploit homologous recombinat

In order to make a directed genetic change in a specific mouse gene we exploit homologous recombinat

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In order to make a directed genetic change in a specific mouse gene we exploit homologous recombination just as we have discussed for E. coli and S. cerevisiae . However, this is much harder to do in mammalian cells than bacteria and yeast. In yeast, when a linear DNA duplex is introduced into the cell, about 90% of the time that that DNA is integrated into the yeast genome it is done by the homologous recombination machinery such that incoming DNA fragment is swapped for the endogenous gene. In mammalian cells the DNA that is integrated into the genome is almost always at a non-homologous site, and the frequency of homologous replacement of an endogenous sequence is
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Unformatted text preview: about 10-3 to 10-5 . What this means is that we have to allow thousands of integration events to take place, and to be able to identify the integration event we wantnamely an integration even that took place by homologous recombination. The first crucial development for this technology was being able to grow the totipotent cells from preimplantation embryos in culture in the lab; these are called mouse embryonic stem cells (ES cells); the crucial development was to devise a clever way to select integrated a DNA construct by homologous recombination . Images removed due to copyright reasons. Cells...
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This note was uploaded on 11/05/2011 for the course BIOLOGY MCB2010 taught by Professor Jessicadigirolamo during the Fall '10 term at Broward College.

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