The NINJA (iNversion Inducible Joined neoAntigen) mouse model was designed to eliminate both the leaky expression of neoantigens in the thymus and the potential for the development of tolerance in developing CD4 and CD8 T cells. NINJA uses a combination of RNA splicing, DNA recombination and three levels of regulation to eliminate leakiness and provide tight control of neoantigen expression. Mice homozygous for the NINJA allele are viable and fertile. The NINJA Gt(ROSA)26 knock-in construct is divided into a regulatory module that controls expression of a fused flpo/ERT2 and a neoantigen module that is a flpo-inducible neoantigen derived from lymphocytic meningitis virus (LCMV) contained within a GFP reporter. The two modules are separated by an insulator sequence that blocks the CAG enhancer from activating tetO. The neoantigen expresses peptides from the LCMV glycoprotein, which are recognized by H2-Db-restricted CD8 T cells (GP-33-41) and H2-I-Ab restricted CD4 T cells (GP67-77). Specifically, the regulatory portion of the vector carries a loxP-flanked flpo fused to human ERT2 under the control of the doxycycline-inducible tetO promoter. The flpo/ERT2 sequence has been spliced and split into 2 exons placed in opposite orientations divided by a polyA. Cre recombinase-mediated inversion is required for a functional protein. Following cre recombination, the regulator module is in a &ldquo;poised&rdquo; state and requires an rtTA/tTA allele and activation with doxycycline/tamoxifen for flpo expression.The neoantigen portion of the vector (inserted in reverse orientation to the regulatory portion) contains a CAG promoter directing expression of the N-terminal portion of GFP followed by an FRT-flanked neoantigen with FLAG-TAG epitope and the C-terminal portion of GFP. Flpo expression results in a permanent inversion of exon 2 of the neoantigen, which lines up splicing sites and allows transcription of the full-length GFP containing the peptide antigen. When activated, GFP+ cells express the neoantigen constitutively and are
 susceptible to recognition by endogenous GP33-41 specific CD8 T cells 
and GP66-77 specific CD4 T cells. This strain may be useful for studies 
of how T cells respond to defined neoantigens in the context of 
peripheral tolerance, autoimmune diseases, and developing tumors.

The use of this mouse model is subject to the terms and conditions of the Limited Use Label License from Caribou Biosciences, Inc.

The use of this mouse strain or other product (the "Licensed Product") is subject to the terms and conditions of the Limited License from The Broad Institute.

The NINJA (iNversion Inducible Joined neoAntigen) mouse model was designed to eliminate leaky expression of neoantigens in the thymus by allowing inducible expression of an LCMV-derived neoantigen following induction. Induction is controlled by a regulatory module containing a cre/tamoxifen/doxycycline-inducible flpo/ERT2, which acts on the neoantigen module containing a flpo-induced neoantigen within a GFP reporter. When activated, GFP+ cells express the neoantigen constitutively and are susceptible to recognition by endogenous GP33-41 specific CD8 T cells and GP66-77 specific CD4 T cells. This strain may be useful for studies of how T cells respond to defined neoantigens in the context of peripheral tolerance, autoimmune diseases, and developing tumors.

This strain is hosted by NIH's MMRRC, which partners with JAX for the distribution of this and other strains. For additional information and to purchase, please visit the MMRRC site.