Fig. 4

CRISPR/Cas9-mediated targeting of Bmp5 leads to fewer ampullary organs developing. Sterlet crispants at stage 45 after in situ hybridisation (ISH) for the hair cell and electroreceptor marker Cacna1d (also expressed in taste buds on the barbels) or the electroreceptor-specific marker Kcnab3. All crispants shown are from the same batch of siblings/half-siblings (in vitro fertilisation used a mix of sperm from three different males). Black arrowheads indicate examples of neuromasts; white arrowheads indicate examples of ampullary organs. Crispants are numbered for cross-referencing with data provided for each crispant in Supplementary file 2. (A,B) In a control Tyr crispant, Cacna1d expression shows the normal pattern of neuromast lines flanked by fields of ampullary organs. The higher power view shows the posterior preopercular ampullary organ field. (C?F) In Bmp5 crispants, Cacna1d expression reveals fewer ampullary organs (compare C,E with A); this phenotype is particularly prominent in the posterior preopercular ampullary organ field (compare D,F with B). (G?I) In a control Tyr crispant, electroreceptor-specific Kcnab3 expression shows the normal distribution of ampullary organs. (J?L1) In a Bmp5 crispant, Kcnab3 expression shows fewer ampullary organs (compare J-L with G-I). Post-ISH Sox2 immunostaining for supporting cells (J1,K1,L1) demonstrates that neuromasts have formed normally. Very few "additional" ampullary organs appeared (i.e., Sox2-positive, Kcnab3-negative ampullary organs: compare J1,K1,L1 with I,J,K); examples are indicated with indented white arrowheads. (Non-lateral line Sox2 expression is also seen in gill filaments and in taste buds on the barbels and around the mouth.) Abbreviations: di, dorsal infraorbital ampullary organ field; ds, dorsal supraorbital ampullary organ field; e, eye; ppp, posterior preopercular ampullary organ field; S, stage; vi, ventral infraorbital ampullary organ field; vs, ventral supraorbital ampullary organ field. Scale bar: 250 ?m.