Gene-targeting strategy of the Cre gene into the Cebpa locus and analysis of cre expression in various organs using the R26 lacZ and R26 EYFP reporter mouse. (A) (1) Genomic structure of the single exon Cebpa locus; the coding sequence is indicated in black, (2) the cebpa-Cre targeting vector, (3) the targeted allele, (4) the targeted allele with deletion of the puromycin cassette (puroδ). Black triangle represents Frt sequence. Ba indicates BamHI; Bg, BglII; E, EcoRI; H, HindIII; Na, NaeI; and Nc, NcoI. (B) Southern blot analysis with 5′probe on EcoRI-digested DNA, with 3′probe on BglII-digested DNA, or with cre-probe on HindIII-digested DNA from PCR⁺ ES clones. (C) Nested PCR on the 3′ homology arm of the Cebpa locus on DNA from puromycin-resistant ES clones (left panel). PCR on DNA from Cebpa^cre/+ mice crossed with FLPeR deleter mice with primers Cre seq F1 and Cre ert2 R showing germline transmission (middle panel), and with primers Cre seq F1 and cebpa R1 showing deletion of the puromycin cassette (right panel). (D) Crossing Cebpa^cre/+ mice with R26 lacZ reporter mice reveals Cebpa-driven cre expression in liver (left) and lungs (right) as expected. Images were captured with a Leica DMLB microscope and 10×/0.25 objective, DFC 420, and processed with Leica Application Suite Version 2.7.1R1. (E) Analysis of the percentage of EYFP⁺ cells among peripheral blood leukocyte subsets in Cebpa^cre/+ R26 EYFP mice (mean ± SD of at least 6 mice analyzed). CD11b⁺Gr1⁺ granulocytes, CD11b⁺ CD115⁺ monocytes, and CD49b⁺ NK cells differ significantly from other subsets (***P < .001).