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. 2017 Dec 14;6:e32417. doi: 10.7554/eLife.32417

Figure 2. Generation of a Salsa6f transgenic mouse line targeted to the Rosa26 locus.

Figure 2.

(A) Transgenic targeting vector for Salsa6f, inserted between Rosa26 homology arms and electroporated into embryonic stem cells. CAG Pr: cytomegalovirus early enhancer/chicken β-actin promoter; Salsa6f: tdTomato-V5-GCaMP6f; FRT, LoxP, AttB, AttP: recombinase sites; WPRE: woodchuck hepatitis virus post-transcriptional regulatory element; pA: bovine growth hormone polyadenylation sequence; NeoR: neomycin resistance gene. (B) Correctly targeted ES cells were screened by Southern blot after HindIII digest for the 5’ end (top) or BglI digest for the 3’ end (bottom). The two clones marked in red failed to integrate at the 5’ end. (C) PCR screening for chimeras based on presence of the Nnt mutation, present only in JM8.N4 ES cells but not in the C57BL/6J blastocyst donors. 2540 and 2543 are chimeras. Control lanes on the right are wild type (Nnt+/+), heterozygous (Nnt+/-), or homozygous mutant (Nnt-/-).