Development and Application of a KASP-Based SNP Marker System for Fingerprinting, and Genetic Diversity Analysis in Tomato

Tomato is an important vegetable crop worldwide with diverse uses. However, long-term domestication and modern breeding have narrowed the genetic basis of mainstream cultivars, leading to high genetic similarity among varieties. Therefore, developing practical molecular markers through genomic approaches has become an important way to effectively identify and utilize germplasm resources and overcome current breeding bottlenecks. This study aimed to screen high-quality single-nucleotide polymorphism (SNP) loci, develop reliable competitive allele-specific PCR (KASP) markers for genotyping tomato germplasm, and further construct DNA fingerprints and perform genetic diversity analysis, ultimately achieving accurate identification of tomato germplasm. Based on whole-genome resequencing of 241 tomato accessions (inbred lines and wild germplasm), 90 were successfully converted into functional KASP markers. According to polymorphic information content (PIC) and minor allele frequency (MAF), 50 core markers were further screened, with an average PIC of 0.357, an average gene diversity (GD) of 0.46, and an average heterozygosity of 0.063. 16 core SNP markers evenly distributed across 12 chromosomes were optimized, which effectively distinguished all tested accessions. Population structure analysis, principal component analysis (PCA), and phylogenetic analysis indicated that the 241 accessions could be clustered into four major subpopulations, consistent with the results derived from whole-genome SNP data. Finally, a DNA fingerprint of the 241 tomato accessions was constructed based on the 16 core markers. This study provides a robust and cost-effective molecular tool for tomato accession identification, germplasm resource conservation, and genetic diversity assessment.