Map-based cloning revealed BhAPRR2 gene regulating the black peel formation of mature fruit in wax gourd (Benincasa hispida).

Map-based cloning revealed BhAPRR2, encoding a two-component response-regulating protein that regulates the black peel formation of mature fruit in wax gourd. Wax gourd is an economically significant vegetable crop, and peel color is a crucial agronomic trait that influences its commercial value. Although genes controlling light green or white peel have been cloned in wax gourd, the genetic basis and molecular mechanism underlying black peel remain unclear. Here, we confirmed that the peel color of wax gourd is a qualitative trait governed by single gene, with black being dominant over green. Through bulked segregant analysis sequencing (BSA-seq) and map-based cloning, we identified Bh.pf3chr5g483 as the candidate gene. This gene encodes a two-component response-regulating protein and is homologous to APRR2, referred to as BhAPRR2. Compared to P170, the BhAPRR2 in YD1 exhibits multiple mutations in both its coding and promoter regions. Notably, the mutations in the coding region do not affect its nuclear localization or transcriptional activation activity. However, the mutations in the promoter region substantially increase its expression in the peel of YD1, potentially contributing to the black peel phenotype observed in this variety. Furthermore, we developed an insertion/deletion (InDel) marker based on a 93-base pair (bp) insertion/deletion mutation in the promoter region of BhAPRR2, which achieved up to 95.8% phenotypic accuracy in a natural population comprising 165 wax gourd germplasms. In summary, our findings suggest that mutations in the promoter region of BhAPRR2 may contribute to the development of black peel in wax gourd. This discovery provides new insights into the molecular and genetic mechanisms underlying peel color diversity and offers a valuable molecular marker for wax gourd breeding efforts.