Abstract

Pumpkin (Cucurbita spp.) is a globally significant vegetable crop known for its high nutritional value and remarkable phenotypic diversity. Yet, the surge in new cultivar releases has overwhelmed traditional morphological descriptors, creating critical gaps in variety purity control and breeders’ rights enforcement. Despite the established utility of SNP markers as the gold standard for genetic analysis, a dedicated high-resolution molecular database for modern pumpkin cultivars remains unavailable. To address this gap, we conducted whole-genome resequencing (WGS) on 94 representative pumpkin cultivars (spanning C. moschata, C. maxima, and C. pepo). Clean reads were mapped to the Cucurbita maxima reference genome. We employed a stringent pipeline to identify genomic variants and utilized STRUCTURE software, Principal Component Analysis (PCA), and Neighbor-Joining (NJ) trees to evaluate population stratification. Linkage disequilibrium (LD) decay and DNA fingerprinting barcodes were also developed. A total of 8,873,150 high-quality variants were identified, including 7,345,007 SNPs and 1,528,143 InDels, with an average SNP density of 21,281.50 SNPs/Mb. Population analysis consistently categorized the 94 cultivars into two primary subpopulations (G1 and G2). The first two PCs accounted for 74.06% of the total genetic variance. Further analysis revealed that G1 possessed a more complex genetic architecture and slower LD decay compared to G2, suggesting distinct selection histories. Finally, we screened for highly informative biallelic SNPs to construct a DNA fingerprinting database, enabling precise sample discrimination through unique chromatic barcodes. This study fills a critical gap in pumpkin genomics by establishing a high-density SNP database and a robust fingerprinting system. These resources provide a definitive tool for variety certification, seed purity testing, and the advancement of molecular-assisted breeding in pumpkin.

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