Phenotypic polymorphisms can be applied to study the micro‐evolutionary forces that maintain genetic variation and can mediate speciation, but it can be difficult to determine the genetic basis of polymorphisms. Recently, restriction‐site‐associated DNA (RAD) sequencing has become popular, which can easily produce multiple single nucleotide polymorphisms from whole genomes. Here, we combined RAD sequencing, allele‐specific PCR and Sanger sequencing to determine the genetic basis underlying male colour dimorphism of a Lake Tanganyika cichlid fish, Cyprichromis leptosoma. Our analyses using both a cross‐family (two parents and 12 F2 males) and 64 wild individuals do not contradict a hypothesis that two alleles of one‐locus control male colour dimorphism. Also, the locus may be located on a genome region that experiences reduced levels of recombination. Although more analyses will be needed to conclude these findings, this study is the first to suggest the genetic basis of a colour polymorphism using RAD sequencing.