miR-144 targets klfd in physiologic context. (A,B) Expression of EGFP mRNAs and EGFP fluorescence in 16 and 24 hpf F2 embryos derived from either Tg(zgata-1:EGFP) transgenic line 2 (A) or Tg(zgata-1:EGFP-klfd 3′-UTR) transgenic line 5 (B). Arrowheads indicate nonspecific expression of EGFP mRNAs in the dorsal neurons. (C) Western blot analyses of EGFP protein in 16 and 24 hpf embryos from either Tg(zgata-1:EGFP) transgenic line 2 or Tg(zgata-1:EGFP-klfd 3′-UTR) transgenic line 5. (D,E) Rescue of EGFP mRNAs by microinjection of either miR-144 or control LNA into 1 cell–stage F2 embryos from Tg(zgata-1:EGFP-klfd 3′-UTR) transgenic line 5. (F) Western blot analysis of endogenous Klfd protein levels in miR-144-deficient embryos at 22 hpf.