Bitter gourd is an economically important vegetable and medicinal crop distinguished by its bitter fruits. Its stigma color is widely used to assess the distinctiveness, uniformity, and stability of bitter gourd varieties. However, limited researches have been dedicated to genetic basis of its stigma color. In this study, we employed bulked segregant analysis (BSA) sequencing to identify a single dominant locusMcSTC1located on pseudochromosome 6 through genetic mapping of an F2population (n=241) derived from the cross between green and yellow stigma parental lines. An F2-derived F3segregation population (n= 847) was further adopted for fine mapping, which delimited theMcSTC1locus to a 13.87 kb region containing one predicted geneMcAPRR2(Mc06g1638), a homolog of theArabidopsistwo-component response regulator-like geneAtAPRR2. Sequence alignment analysis ofMcAPRR2revealed that a 15 bp insertion at exon 9 results in a truncated GLK domain of its encoded protein, which existed in 19 bitter gourd varieties with yellow stigma. A genome-wide synteny search of the bitter gourdMcAPRR2genes in Cucurbitaceae family revealed its close relationship with other cucurbits APRR2 genes that are corresponding to white or light green fruit skin. Our findings provide insights into the molecular marker-assisted breeding of bitter gourd stigma color and the mechanism of gene regulation for stigma color.