Direct removal in the mouse of a flexed neo gene from a three-loxP conditional knockout allele by two novel approaches

The presence in an intron of the ploxP-neo-loxP cassette often results in s evere interference with gene expression. Consequently, many investigators s electively remove the ploxP-neo-loxP cassette by transient expression of Cr e in ES cells, Although effective, the added manipulation of the ES cells m ay reduce the likelihood that a clone will be able to transmit via the germ line, Therefore, we developed two novel approaches that remove the ploxP-ne o-loxP by Cre-mediated recombination in mouse. First, the ploxP-neo-loxP-co ntaining mice were crossed with Ella-Cre transgenic mice. Second, a Cre-exp ression plasmid was injected into pronuclei of fertilized eggs bearing the ploxP-neo-loxP allele, Both approaches produced mosaic mice with partial an d complete excision. These mosaic mice were then mated, and the neo-less co nditional 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. genesis 30: 1-6, 2001, Published 2001 Wiley-Liss, Inc.(dagger)