Direct removal in the mouse of a floxed neo gene from a three-loxP conditional knockout allele by two novel approaches |
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Authors: | Xu X Li C Garrett-Beal L Larson D Wynshaw-Boris A Deng C X |
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Institution: | Genetics of Development and Disease Branch, National Institute of Diabetes, Digestive and Kidney Diseases, Bethesda, Maryland, USA. |
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Abstract: | The presence in an intron of the ploxP-neo-loxP cassette often results in severe interference with gene expression. Consequently, many investigators selectively remove the ploxP-neo-loxP cassette by transient expression of Cre in ES cells. Although effective, the added manipulation of the ES cells may reduce the likelihood that a clone will be able to transmit via the germline. Therefore, we developed two novel approaches that remove the ploxP-neo-loxP by Cre-mediated recombination in mouse. First, the ploxP-neo-loxP-containing mice were crossed with EIIa-Cre transgenic mice. Second, a Cre-expression plasmid was injected into pronuclei of fertilized eggs bearing the ploxP-neo-loxP allele. Both approaches produced mosaic mice with partial and complete excision. These mosaic mice were then mated, and the neo-less conditional knockout allele was found in the offspring after screening only a few litters. These procedures provide options for removing neo directly in the mouse in addition to the commonly used approach that deletes neo in ES cells. |
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Keywords: | conditional knockout neo deletion Brca1 EIIa‐Cre pronuclear injection |
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