Fig 6. Transient puromycin selection results in high KI efficiency of fluorescence reporter genes into mESCs.

(A, B, C) PCR tests for fluorescent reporter KI at the indicated loci in mESCs. For A and B, primers amplifying a region inside the KI gene and outside the donor plasmid template were used. For C, primers spanning the integration region were used to distinguish between homozygous and heterozygous clones. Expected amplicon sizes are shown. WT = wildtype. For KI assessment in B, lanes 9 and 17 were not counted as positive KI because the amplicon did not run at the predicted size. (D) Phase and fluorescence microscopy of a homozygous Six6-P2A-eGFP reporter KI line generated in C. mESCs were differentiated to optic vesicles for 8 days. Scale bar = 275 μm.