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Figure S4. Control experiments for setd7-MO knockdown approach in Xenopus embryos.
(A) Whole mount ISH analysis of ptf1a and pdx1 in uninjected (ctrl), 5bp-mismatched setd7
morpholino oligonucleotide (setd7-5MM MO)-injected (5 ng) and setd7-MO-injected (5 ng)
Xenopus embryos. A 5bp-mismatched setd7 MO was designed introducing 5 G to C or C to G
substitutions distributed evenly through the sequence and used as additional specificity control in
the knockdown experiments (Eisen, Smith, 2008) (see Material and Methods for the sequences).
While injection of 5 ng of setd7-MO led to strong reduction of pancreatic genes, the equivalent
dose of setd7-5MM MO did not perturb pancreas formation, providing further evidence of
morpholino specificity. Arrowheads indicate dorsal and ventral pancreatic buds. Brackets indicate
ptf1a expression in the eye and hindbrain. Size bar, 1mm. n=3. (B) Whole mount ISH analysis
using ptf1a and insulin antisense probes. Tadpole embryos (st. 36) injected with setd7-MO
showed reduction of ptf1a and insulin expression domains (see arrowheads; 70%). ptf1a and
insulin expression was rescued in embryos injected with setd7-MO and mouse Setd7 (500 pg)
mRNA (~70-80%). Embryos left untreated (ctrl) show normal ptf1a expression in both pancreatic
buds, as indicated by the arrowheads, and hindbrain (white bracket), as well as insulin expression
in the dp (yellow arrowheads). n=3.
Development • Supplementary information |
