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Background: Genetic markers are crucial for breeding crops with desired agronomic traits, and their development can be expedited using next-generation sequencing (NGS) and bioinformatics tools. Numerous tools have been developed to design molecular markers, enhancing the convenience, accuracy, and efficiency of molecular breeding. However, these tools primarily focus on genetic variants within short user-input sequences, despite the availability of extensive omics data for genomic variants. To design molecular markers encompassing a vast number of genetic variants at the genome-wide scale in soybean, an automatic system capable of handling NGS-based big data is necessary. Results: In this study, we developed a robust digital platform, the CAPS Maker, for designing cleaved amplified polymorphic sequence (CAPS)/derived CAPS (dCAPS) markers in soybeans. This platform simplifies the systematic design of genomic markers with a user-friendly graphical interface, featuring a 'SNP Browser' and 'Primer Table', along with internal programs (e.g., the eHT-PCR module) to design unique primer pairs for highly duplicated genomes like soybean. Conclusions: The CAPS Maker's efficiency and reliability were experimentally verified by comparing its marker predictions with actual experimental results. Consequently, breeders can easily design CAPS/dCAPS markers using the CAPS Maker platform to develop new soybean cultivars with beneficial agronomic traits. This platform is freely accessible at https://tgil.donga.ac.kr/CAPSMaker. [ABSTRACT FROM AUTHOR]

The soybean is agro-economically the most important among all cultivated legume crops, and its seed color is considered one of the most attractive factors in the selection-by-breeders. Thus, genome-wide identification of genes and loci associated with seed colors is critical for the precision breeding of crop soybeans. To dissect seed pigmentation-associated genomic loci and genes, we employed dual approaches by combining reference-based genome-wide association study (rbGWAS) and k-mer-based reference-free GWAS (rfGWAS) with 438 Glycine accessions. The dual analytical strategy allowed us to identify four major genomic loci (designated as SP1-SP4 in this study) associated with the seed colors of soybeans. The k-mer analysis enabled us to find an important recombination event that occurred between subtilisin and I-cluster B in the soybean genome, which could describe a special structural feature of ii allele within the I locus (SP3). Importantly, mapping analyses of both mRNAs and small RNAs allowed us to reveal that the subtilisin-CHS1/CHS3 chimeric transcripts generate and act as an initiator towards 'mirtron (i.e., intron-harboring miRNA precursor)'-triggered silencing of chalcone synthase (CHS) genes. Consequently, the results led us to propose a working model of 'mirtron-triggered gene silencing (MTGS)' to elucidate a long-standing puzzle in the genome-wide CHS gene silencing mechanism. In summary, our study reports four major genomic loci, lists of key genes and genome-wide variations that are associated with seed pigmentation in soybeans. In addition, we propose that the MTGS mechanism plays a crucial role in the genome-wide silencing of CHS genes, thereby suggesting a clue to currently predominant soybean cultivars with the yellow seed coat. Finally, this study will provide a broad insight into the interactions and correlations among seed color-associated genes and loci within the context of anthocyanin biosynthetic pathways. [ABSTRACT FROM AUTHOR]