Effectiveness of RPS19 morpholinos and hemoglobin staining of embryos coinjected with Rps19 mRNA and P53 MO using o-dianisidine.

The Rps19:egfp construct was made by inserting a partial sequence of Rps19 cDNA (containing 60 bps from the 52 UTR ) with the N-terminus of egfp into modified pEGFP-N1 (the ATG codon of EGFP was removed). The sequence of RPS19 MO1 is a compliment of bp 1–24 of Rps19 cDNA. Embryos co-injected with 25 ng Rps19:egfp DNA and 5 ng control MO produced green fluorescent fusion protein (A), and expression of the fusion protein was inhibited by co-injection with 2 ng Rps19 Mo (B). O-staining results show a drastic reduction in the number of hemoglobin-stained blood cells when Rps19 is knockdown (C and D are the control, E and F are Rps19 knockdown) and partially reversed by co-injection of P53 morpholino (G and H). A, B, D, F and H are the lateral view; C, E and G are the ventral view.

RPS19 is not required for HSC formation and RPS19 knockdown embryos have normal number of gata1-positive cells. (A–C) The expression of cmyb (black arrow) was comparable in RPS19 morphants and RPS19 and P53 double morphants at 48 hpf. (D–F) The expression of runx1 was comparable in RPS19 morphants and RPS19 and P53 double morphants at 48 hpf. (G–I) The expression of gata1 was comparable in RPS19 morphants and RPS19 and P53 double morphants at 24 hpf. A–I is lateral view.

The in situ results of fzd5, alas2, nrp1a and tagln in zebrafish embryos at 48hpf. (A–C) The expression of fzd5 (black arrow) in control, RPS19 morphants and RPS19 and P53 double morphants at 48 hpf. (D–F) The expression of alas2 (black arrow) in control, RPS19 morphants and RPS19 and P53 double morphants at 48 hpf. (G–I) The expression of nrp1a (black arrow) in control, RPS19 morphants and RPS19 and P53 double morphants at 48 hpf. (J–L) The expression of tagln (black arrow) in control, RPS19 morphants and RPS19 and P53 double morphants at 48 hpf. A-L is lateral view.