Extended Data Figure 1 |. Design and characterization of Rosa26^uLIPSTIC mice.

(A) The uLIPSTIC cassette carrying the lox-stop-lox G₅Thy1.1 followed by mSrtA-PDGFRtm fused to FLAG tag was cloned into the Ai9 Rosa26 targeting plasmid. (B) Insertion of the uLIPSTIC cassette was assessed in embryonic stem (ES) cells by Southern blotting using a ³²P-labeled probe (Supplementary Table 8 and Supplementary Figure 1 for gel source data) annealing upstream of the left arm after EcoRI digestion. ESCs carrying the insertion exhibit an extra EcoRI restriction site, resulting in a 7.4 kb fragment upon enzymatic digestion. The blot shows 2 heterozygous integrations out of 7 ES cell clones screened. (C-E) The specificity and efficiency of Rosa26^uLIPSTIC recombination are determined by the Cre driver used. (C) Representative gating strategy for resident dendritic cells (rDCs; LIN^–, MHC-II^int, CD11c^hi), migratory dendritic cells (mDCs; LIN^–, MHC-II^hi, CD11c⁺), CD4⁺ T cells, CD8⁺ T cells, regulatory T (Treg) cells and B cells in lymph nodes. (D) SrtA expression (determined by FLAG detection) is induced by Cre recombination. Use of a constitutive Cre line (e.g., CD4-Cre) results in efficient but non-specific SrtA expression, generating T cells that can only be used in adoptive cell transfer experiments. The use of inducible Cre lines such as CD4-CreERT2 and Foxp3^CreERT2 can often resolve specificity issues, enabling the implementation of uLIPSTIC in fully endogenous models. (E) SrtA expression in conventional DC subsets 1 (cDC1) and 2 (cDC2) in Rosa26^uLIPSTIC/WT.Clec9a^Cre/WT mice.