Figure 4

Tracing of gonad development in pld6 mutant. A) Comparison of PGC numbers between wildtype and mutant by WISH with vasa probe. WISH was performed on embryos at 24, 48, 72, and 96 hpf. N represents analyzed embryo number. B) Statistical analysis of total PGC numbers in wildtype and pld6 ^−/− embryos. C) RT‐qPCR detection of the expression of germ plasm factors (vasa, piwil1, buc, and nanos3) in wildtype and pld6 ^−/− larvae. The decreased expression level of pld6 was set as positive control. D–F) Tracing of germ cell development in wildtype and pld6 ^−/‐ juvenile gonads at 22 dpf (D), 30 dpf (E), and 45 dpf (F). Anti‐vasa staining was performed to label the germ cells, and DAPI staining was performed to label the nuclear. Green arrows marked the stage IA and IB oocytes. N represents analyzed individual number. Scale bar: 100 µm. G) Anti‐Nanos2 immunostaining of wildtype and mutant gonads at 30 dpf. N represents analyzed individual number. Scale bar: 20 µm. The data were expressed as mean ± SD. The P values in this figure were calculated by two‐sided t‐test. *P < 0.05; ***P < 0.001; ns, no significant difference; PGC, primordial germ cell; WISH, whole‐mount in situ hybridization; hpf, hours post fertilization; dpf, days post fertilization; RT‐qPCR, reverse‐transcription quantitative PCR.