Dissecting seed pigmentation-associated genomic loci and genes by employing dual approaches of reference-based and k-mer-based GWAS with 438 Glycine accessions

Jin-Hyun Kim,Ho-Won Jung,Joo-Seok Park,Chae-Young Lee,Min-Gyun Jeong,Hong-Kyu Choi,Yongsoo Choi,Jiu Liang Xu,Istvan Rajcan

doi:10.1371/journal.pone.0243085

Abstract

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.

Highlights

The soybean [Glycine max (L.) Merr.] is the most agro-economically important among all domesticated legume crops
Even if a certain genome is perfectly assembled as a standard framework for the reference, reference-based genome-wide association study (rbGWAS) may retain their innate limitations against calling every single variation, mainly due to the genome’s complexity and large scale genomic changes unaccounted in common Genome-wide association study (GWAS) analyses, all of which may cause the ‘missing heritability’ problem [21]
To precisely identify and dissect genes and genomic loci associated with the pigmentation of soybean seeds, this study employed an integrated approach in both analytical means and data resources

Summary

Introduction

The soybean [Glycine max (L.) Merr.] is the most agro-economically important among all domesticated legume crops. In addition to its nutritional value, the soybean contains a variety of bioactive phytochemicals such as saponin, lecithin, isoflavone, anthocyanin and many others [2]. Anthocyanin is one of the key components that exert antioxidant effects and determine the seed colors of soybean. The seed color should be considered one of the most critical qualitative traits for grain crops like soybeans. The seed coat of the soybean can be represented by four colors; black, brown, yellow, green, and mottled. Black soybeans are recently attracting more interest due to their antioxidant properties and flavors [6]

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Conclusion