Supplemental Material for Kottler et al., 2019 KottlerVerena A. FeronRomain NandaIndrajit KloppChristophe DuKang KneitzSusanne HelmprobstFrederik LamatschDunja K. Lopez-RoquesCéline LluchJerôme JournotLaurent ParrinelloHugues GuiguenYann SchartlManfred 2019 Figure S1 presents the results of the RAD-sequencing analysis. Figure S2 shows the analysis of <i>G. affinis </i>and <i>G. holbrooki </i>pooled-sequencing reads aligned to the genome of <i>X. maculatus </i>using 10 kb windows. Figure S3 represents a molecular phylogeny of fish <i>sox9</i>. Figure S4 are alignments of male <i>G. holbrooki </i>and <i>G. affinis </i><i>gipc1</i>and <i>sox9b </i>alleles and protein sequences. Figure S5 are multiple species alignments of Gipc1 and Sox9 protein sequences. Figure S6 contains PCR results demonstrating that <i>sox9bY </i>was generated by allelic diversification. Figure S7 contains pictures of melanic and non-melanic <i>G. holbrooki</i>. Table S1 gives information on the fish used for pool sequencing. Table S2 presents the results of the allele-specific PCRs. Table S3 contains the annotated loci within the 100 kb windows with the highest density of sex-specific SNPs. Table S4 contains the sex chromosome genotypes of F<sub>1 </sub>fish (pedigree WLC20074) from a cross between a <i>G. affinis </i>female (WLC1313) and a <i>G. holbrooki </i>male (WLC2514) determined by PCR. Table S5 gives information on the number of offspring of non-melanic females and non-melanic males derived from melanic strains.