Gonads were collected from zebra finch embryos at Hamburger Hamilton stage 28. Seven male and seven female embryonic gonads were pooled in 100 ul trypsin 0.05% with same-sex and incubated at 37℃ for 5 min with gentle pipetting. The digestion was stopped with 100 ul of Dulbecco's Modified Eagle Medium containing 10% fetal bovine serum and washed with 500 ul PBS. Cells were pelleted down and resuspended in PBS. Single cell libraries were prepared using the Chromium controller according to the 10x chromium Next GEM Single Cell 3' v3.1 protocol. Libraries were prepared using Chromium Next GEM Single Cell 3p RNA library v3.1. Single Cell 3' GEX and feature barcode libraries are sequenced on the Illumina sequencing system.