Gt(ROSA)26Sor^{tm1.2(CAG-COX4I1/APX1*)Ddg}

A CAG promoter, FRT-flanked Stop cassette, loxP-flanked Stop cassette (in reverse transcriptional orientation), Matrix-dAPEX2, WPRE, bGHpA, attB, PGK-Neo, PGKpA, and attP were introduced to intron 1 via homologous recombination. APEX2 is an enhanced (K14D, W41F, E112K, A134P) soybean ascorbate peroxidase (APX1) that is less catalytically active than horseradish peroxidase (HRP), but is functional when expressed in cytosolic environments, unlike HRP. Dimeric APEX2 (dAPEX2) contains the reversion of K14 and E112 and is presumed to enhance heme affinity and thermal stability of the protein. Targeting of dAPEX2 to the mitochondrial matrix to create Matrix-dAPEX2 was accomplished by a targeting peptide from human COX4I1 (MLATRVFSLVGKRAISTSVCVRAH) followed by a linker sequence (KDP) and V5 epitope tag. Matrix-dAPEX2 has the same sequence as mito-V5-APEX2 (Addgene plasmid 117176) except for the replacement of APEX2 with dAPEX2. Cre-mediated recombination removed the floxed stop. The FRT-flanked stop cassette remains intact. (J:280094)