Metrics of sequence constraint overlook regulatory sequences in an exhaustive analysis at phox2b

In situ hybridization (ISH) of endogenous phox2b expression. ISH was performed on wild-type zebrafish embryos from 24 to 96 hpf using a dig-labeled phox2b RNA probe. (a) Dorsal view of 24-hpf embryo, illustrating phox2b expression in the hindbrain and anterior spinal column. (b) Dorsal view of 48-hpf embryo. hb, hindbrain; mo, medulla oblongata; ventral diencephalon (filled arrowhead), locus coeruleus (open arrowhead). (c) Lateral view of 48-hpf embryo. Rhombomeres of the hindbrain are numbered; mo, medulla oblongata; locus coeruleus (open arrowhead); cranial ganglia (black arrow). (d) Lateral view of the trunk of a 96-hpf embryo. Spinal cord (open dotted arrow) and ENS (open arrows).