The Notch1 activity-trap knock-in allele, N1IP::Cre^HI, has the endogenous Notch1 intracellular domain (NICD1) replaced by a Cre recombinase with nuclear localization signal. Like endogenous Notch1, the Notch1-Cre fusion protein is expressed on the cell membrane and its extracellular domain binds Notch ligands (-Delta, Serrate and Lag-2), but proteolytic cleavage of the transmembrane domain tether results in release of nuclear-localized Cre rather than NICD1. Cre then enters the nucleus, where it results in the excision of floxed sequences. If a Cre-dependent reporter is present, Cre recombinase activity will 'trap' the cells and permanently mark its descendent lineage independently of any further Notch1 activation. Specific cell types are described below.
Heterozygous mice are viable and fertile with no gross phenotypic abnormalities. Homozygotes die at embryonic day 9.5 (E9.5).

The Notch1 activation-dependent reporter knock-in alleles, N1IP::Cre^LO (Stock No. 006953), N1IP::Cre^ERT2 (Stock No. 027235) and N1IP::Cre^HI (Stock No. 027234) are compared below.

N1IP::Cre^LO (and N1IP::Cre^ERT2 in the presence of tamoxifen) is relatively insensitive and seems to predominantly 'trap' cells in which moderate-to-high levels of sustained Notch activity or repeated activation cycles are known to occur (e.g., endothelium), while missing cells in many tissues known to rely on Notch1 activity (e.g., the hematopoietic system). The donating investigator speculates these differences could be attributed to the reduced transcript stability of the NICD (due to the insertion of 6xMyc-tags at the C-terminus of the nuclear-localized Cre recombinase). Also, in some tissues, sequences required for proper membrane trafficking of Notch may have been lost when NICD1 was replaced with Cre recombinase.

The N1IP::Cre^HI allele lacks the C-terminal 6xMyc-tag and has one more additional polyadenylation signal following the nuclear-localized Cre recombinase. These improvements restore the Cre recombinase C-terminus and increase transcript stability two-fold, respectively; resulting in a cumulative five- to ten-fold increase in Cre activity. The resulting Notch1-Cre fusion protein is cleaved as efficiently as endogenous Notch1, and has significantly improved sensitivity compared to N1IP::Cre^LO. This improved sensitivity allows users to 'trap' cells experiencing moderate-to-lower Notch activation thresholds; including most cells/tissues known to utilize Notch1 during their development and/or maintenance.