Molecular characterization of proton beam-induced mutations in soybean using genotyping-by-sequencing

Proton beam irradiation is a next-generation technique to develop mutant crop varieties. The mutagenic effects and molecular mechanisms of radiation are important multi-disciplinary research subjects. This study was conducted to investigate the types of mutations induced in the soybean genome by proton beam irradiation. In total, 22 plants, including 10 M₂ plants treated with proton beam irradiation at 118 and 239 Gy, each, and two wild-type plants (Daepung) were sequenced by genotyping-by-sequencing (GBS). In total, 7453 single nucleotide polymorphisms (SNPs) were detected in the 20 M₂ plants, compared with the two wild-type controls. The SNP frequency was 1/36,976 bp with proton beam irradiation at 118 Gy, and 1/32,945 bp at 239 Gy. Of these, 3569 SNPs were detected in genic regions. We observed that proton beam irradiation induced more substitutions than small insertion-deletions (INDELs). Based on the mutagenic effect of proton beam irradiation, the frequency of transition mutations was shown to be higher than that of transversions. The proton beam-induced SNPs were distributed uniformly in most of the chromosomes. Gene ontology (GO) analysis showed that there were many genes involved in protein metabolic process under biological process, intracellular membrane-bounded organelle under cellular component, and nucleic acid binding under molecular function. This study could provide valuable information for investigating the potential mechanisms of mutation, and guidance for developing soybeans cultivars using mutation breeding.