Bo-Keun Ha

Development, validation and genetic analysis of a large soybean SNP genotyping array

Cultivated soybean (Glycine max) suffers from a narrow germplasm relative to other crop species, probably because of under-use of wild soybean (Glycine soja) as a breeding resource. Use of a single nucleotide polymorphism (SNP) genotyping array is a promising method for dissecting cultivated and wild germplasms to identify important adaptive genes through high-density genetic mapping and genome-wide association studies. Here we describe a large soybean SNP array for use in diversity analyses, linkage mapping and genome-wide association analyses. More than four million high-quality SNPs identified from high-depth genome re-sequencing of 16 soybean accessions and low-depth genome re-sequencing of 31 soybean accessions were used to select 180,961 SNPs for creation of the Axiom(®) SoyaSNP array. Validation analysis for a set of 222 diverse soybean lines showed that 170,223 markers were of good quality for genotyping. Phylogenetic and allele frequency analyses of the validation set data indicated that accessions showing an intermediate morphology between cultivated and wild soybeans collected in Korea were natural hybrids. More than 90 unanchored scaffolds in the current soybean reference sequence were assigned to chromosomes using this array. Finally, dense average spacing and preferential distribution of the SNPs in gene-rich chromosomal regions suggest that this array may be suitable for genome-wide association studies of soybean germplasm. Taken together, these results suggest that use of this array may be a powerful method for soybean genetic analyses relating to many aspects of soybean breeding.

Genome-wide transcriptome profiling of ROS scavenging and signal transduction pathways in rice (Oryza sativa L.) in response to different types of ionizing radiation.

Ionizing radiation directly and indirectly affects gene expression within the plant genome. To access the antioxidant response of rice to different types of ionizing radiation, rice seeds were exposed to gamma-ray, cosmic-ray and ion beam radiation. Exposure to ionizing radiation dramatically decreased the shoot length in all plants but not the root length compared with a non-irradiated plant. Electron spin resonance, confirmed that the number of free radicals in cell was greatly increased by different types of ionizing radiation. The measurement of the MDA, chlorophyll, carotenoids contents and activity of antioxidant enzymes revealed that gamma-ray and cosmic-ray, but not ion beam, ionization deceased chlorophyll and carotenoids contents, while all three ionization treatments increased the activities of peroxidase, ascorbate peroxidase, and superoxide dismutase compared with the non-irradiated plants. Microarray analysis using Affymetrix GeneChip was used to establish the gene transcript profiles of rice genes regarding ROS scavenging and signal transduction pathways after ionization treatment. Many of the rice genes involved in ROS scavenging and signal transduction pathways showed induction or repression that had increased more than twofold after ionization treatment. In particular, genes associated with electron transport, such as NADPH oxidase-like and alternative oxidase, were often down-regulated by more than twofold in response to the ionization treatments. In our transcriptomic profile analysis, we confirmed that the expression of rice genes associated with ROS scavenging and signal transduction pathways was induced or repressed to different degrees by the different types of ionizing radiations, as in other environmental stresses.

Identification of quantitative trait loci controlling linolenic acid concentration in PI483463 (Glycine soja)

Key messageThe QTLs controlling alpha-linolenic acid concentration from wild soybean were mapped on nine soybean chromosomes with various phenotypic variations. New QTLs for alpha-linolenic acid were detected in wild soybean.AbstractAlpha-linolenic acid (ALA) is a polyunsaturated fatty acid desired in human and animal diets. Some wild soybean (Glycine soja) genotypes are high in ALA. The objective of this study was to identify quantitative trait loci (QTLs) controlling ALA concentration in a wild soybean accession, PI483463. In total, 188 recombinant inbred lines of F5:6, F5:7, and F5:8 generations derived from a cross of wild soybean PI483463 (~15xa0% ALA) and cultivar Hutcheson (~9xa0% ALA) were planted in four environments. Harvested seeds were used to measure fatty acid concentration. Single nucleotide polymorphism markers of the universal soybean linkage panel (USLP 1.0) and simple sequence repeat markers were used for molecular genotyping. Nine putative QTLs were identified that controlled ALA concentration by model-based composite interval mapping and mapped to different soybean chromosomes. The QTLs detected in four environments explained 2.4–7.9xa0% of the total phenotypic variation (PV). Five QTLs, qALA5_3, qALA6_1, qALA14_1, qALA15_1, and qALA17_1, located on chromosomes 5, 6, 14, 15, and 17 were identified by model-based composite interval mapping and composite interval mapping in two individual environments. Among them, qALA6_1 showed the highest contribution to the PV with 10.0–10.2xa0% in two environments. The total detected QTLs for additive and epistatic effects explained 52.4xa0% of the PV for ALA concentration. These findings will provide useful information for understanding genetic structure and marker-assisted breeding programs to increase ALA concentration in seeds derived from wild soybean PI483463.

Identification of candidate genes for an early-maturing soybean mutant by genome resequencing analysis

Flowering is indicative of the transition from vegetative to reproductive phase, a critical event in the life cycle of plants. In this study, we performed whole genome resequencing by Illumina HiSeq to identify changes in flowering genes using an early-flowering phenotype of soybean mutant line Josaengserori (JS) derived from Korean landrace, Seoritae (SR), and we obtained mapped reads of 131,769,690 and 167,669,640xa0bp in JS and SR, respectively. From the whole genome sequencing results between JS and SR, we identified 332,821 polymorphic SNPs and 65,178 indels, respectively. Among these, 30 flowering genes were in SNPs and 25 were in indels. Among 30 flowering genes detected in SNPs, Glyma02g33040, Glyma06g22650, Glyma10g36600, Glyma13g01290, Glyma14g10530, Glyma16g01980, Glyma17g11040, Glyma18g53690, and Glyma20g29300 were non-synonymous substitutions between JS and SR. Changes in Glyma10g36600 (GI), Glya02g33040 (AGL18), Glyma17g11040 (TOC1), and Glyma14g10530 (ELF3) in JS affected the expression of GmFT2a and resulted in early flowering. These results provide insight into the regulatory pathways of flowering in soybean mutants and help to improve our knowledge of soybean mutation breeding.

Alteration of seed storage protein composition in soybean [Glycine max (L.) Merrill] mutant lines induced by γ-irradiation mutagenesis.

This study investigated the alteration of seed storage proteins in soybean mutants induced by γ-irradiation. Five soybean cultivars and four landraces were irradiated with 250 Gy of γ rays to induce variability. The seed storage protein profiles of 414 genetic fixed mutants (M(12)-M(20)) having excellent agricultural traits were analyzed by SDS-PAGE. Among the 414 mutants, 58 were identifed as lacking lipoxygenase, 89 lacking the α subunit, 113 lacking the α subunit, and 40 with an altered β subunit. One hundred and forty-nine mutants lacked the A(3) subunit of glycinin. Fifty-four mutants showed higher trypsin inhibitor (TIA) activity, whereas 139 showed lower TIA activity compared to their original cultivars. The selected mutants with low amounts of antinutritional factors such as trypsin inhibitor, lipoxygenase, and α subunit will constitute genetic resources for improving soybean protein quality.

Comparison of a high oleic acid soybean line to cultivated cultivars for seed yield, protein and oil concentrations

An increased oleic acid concentration improves the nutritional value and oxidative stability of soybean seed oil. High oleic acid soybean cultivars should have desirable agronomic and compositional traits such as a high seed yield, protein and oil concentrations compared to popular, commercially grown cultivars. A high oleic acid soybean line, JD11-0070, was compared to three commercial soybean cultivars in four Korean environments for seed yield, protein and oil concentrations. JD11-0070 had a stable oleic acid concentration across environments, averaging approximately 79xa0% at each location. In addition, there were no significant differences between JD11-0070 and the other cultivars for seed yield or protein concentration, although JD11-0070 had a lower oil concentration than the other cultivars. Thus, high oleic acid soybean lines such as JD11-0070 can be developed with similar yields and favorable seed protein and oil quality compared to commercially grown soybeans. In addition, use of high oleic oil from this line could contribute to human health, and industrial applications.

Assessment of growth and seed oil composition of kenaf (Hibiscus cannabinus L.) germplasm

This study was conducted to evaluate the growth characteristics and fatty acid composition among 15 kenaf mutants derived from the kenaf germplasm C14 and 15 kenaf accessions originating from Russia, India, China, Iran, and Italy. The overall growth performance (plant height, stem diameter, flowering date, leaf, and flower size) of the stem color mutant lines derived from C14 are similar to those of the original variety. However, the flower color mutant lines derived from C14 showed flowering to occur 10 days later when compared with the original variety and showed smaller leaf sizes than the original variety. Late-ripened kenaf accessions (Jinju, Auxu, and Jnagdae) can yield more bio-mass compared with early or medium-maturing germplasm. The late maturity kenaf (Auxu, Jinju, and Jangdae) has a higher oil percentage than the early maturity germplasm. Linoleic, oleic, and palmitic acids were the predominant fatty acids in all kenaf seeds. The stem color mutant lines significantly surpassed the parental means of all saturated fatty acids. In addition, the flower color mutant lines showed broad ranges of variation in oleic acid. The 15 accessions showed a wide range of fatty acid compositions, spanning from 29.75 to 38.30% saturated fatty acids and 61.70 to 70.24% total unsaturated fatty acids, and the late maturity kenaf has a higher linoleic acid percentage than the early maturity germplasm. The flowering period was highly positively (P ≤ 0.01) correlated with the plant height, stem diameter, oil percent, and linolenic acid (C18:3), and it was significantly negatively (P ≤ 0.01) correlated with stearic acid (C18:0). These results will provide valuable information to assist the parental selection of kenaf breeding.

Detection of quantitative trait loci controlling UV-B resistance in soybean

The depletion of stratospheric ozone has resulted in increased amounts of ultraviolet-B radiation (UV-B: 280–320xa0nm) reaching the Earth’s surface. Even a small increase in the incident UV-B radiation could cause significant biological effects in plants. In this study, we identified putative quantitative trait loci (QTL) associated with the resistance to enhanced UV-B radiation in soybean. A population of 115 recombinant inbred lines (RILs) derived from a cross between Keunolkong (a sensitive soybean variety) and Iksan 10 (a resistance soybean line) was analyzed. A total of 110 SSR markers were used to construct a linkage map. RILs and both parents were grown with supplemental UV-B radiation in a greenhouse. In order to screen for UV-B resistance, the degree of leaf chlorosis (DLC), degree of leaf shape change (DLS), degree of petiole color change (DPC) and degree of total plant damage (DTP) were evaluated. Using composite interval mapping analysis, one major QTL associated with all of the characteristics, DLC, DLS, DPC, and DTP, was detected on soybean chromosome 19 between Satt495 and Satt238, accounting for 10.76–32.8xa0% of the phenotypic variance. The ‘Iksan 10’ allele increased the resistance to UV-B radiation for the DLC, DLS, DPC, and DTP traits. This study is the first trial for identification of QTLs associated with UV-B resistance in soybean. In addition, these results provided basic information not only for the improvement of UV-B resistance through marker-assisted selection, but also for the future identification of putative candidate genes of UV-B resistance.

Selection and molecular characterization of a lipoxygenase-free soybean mutant line induced by gamma irradiation

AbstractKey messageA lipoxygenase-free soybean mutant line (H70) induced by gamma ray was selected and its detailed information about the lipoxygenase was analyzed by comparison of DNA sequence.AbstractnSoybean seeds contain three lipoxygenase enzymes, which induce a beany or grassy flavor. The elimination of lipoxygenases can reduce the poor stability and off-flavors of soybean oil and protein products. In this study, we selected a soybean mutant (H70) in which the three lipoxygenases had been mutated using gamma rays. To obtain detailed information about the lipoxygenase, we investigated the sequences of the Lox1, Lox2 and Lox3 genes in H70 compared to the original cultivar, Hwanggum. Comparisons of the sequences of the Lox1 and Lox2 genes in H70 with those in a line with normal lipoxygenase (HG) showed that the mutations in these genes affected a highly conserved group of six histidine residues necessary for enzymatic activity. Lox1 in H70 contained a 74xa0bp deletion in exon 8, creating a stop codon that prematurely terminates translation. A single point mutation (T-A) in exon 8 of Lox2 changed histidine (H532, one of the iron-binding ligands essential for Lox2 activity) to glutamine. The mutation in the Lox3 gene in H70 was a single-point mutation in exon 6 (A-G), which changed the amino acid from histidine to arginine. This amino acid alteration in Lox3 was located in the N-terminal barrel, which might play a role in molecular recognition during catalysis and/or proteolysis. These results suggest that gene analysis based on DNA sequencing could be useful for elucidating the lipoxygenase content in soybean mutant lines. Additionally, the soybean mutant line selected in this study could be used to develop soybean cultivars with improved flavor.

Fruit Quality and Chemical Contents of Hybrid Boysenberry ( Rubus ursinus ) Lines Developed by Hybridization and Gamma Irradiation

The Rubus fruit is an economically important berry crop that contains various functional compounds. The objective of this study was to analyze fruit qualities (i.e., pH, soluble solids content, titratable acidity, and mineral content) as well as fatty acid and phenolic compounds (i.e., ellagic acid and anthocyanins) among hybrid boysenberry lines developed by hybridization and gamma irradiation. There were no significant differences in the hybrid boysenberry fruit pH and titratable acidity (%) among the tested genotypes. However, the soluble solids content was higher in the BSA119 and BSA144 mutants than in the original genotype (BS_Hybrid). Meanwhile, linoleic acid was the most abundant fatty acid in the analyzed hybrid boysenberry fruits. The fatty acid composition did not differ significantly among the genotypes. The ellagic acid content of all genotypes ranged from 8.72 mg/100 g to 46.10 mg/100 g, with the highest concentration observed for the BSB127 genotype. Additionally, cyanidin-3-O-sophoroside (M-H, 611 m/z) and cyanidin-3-O-glucoside (M-H, 449 m/z) were the two major anthocyanins detected in the boysenberry and mutant genotypes, while cyanidin-3-O-glucoside was the predominant blackberry anthocyanin. The total anthocyanin concentrations of four mutant genotypes (i.e., BSA036, BSA078, BSA101, and BSB127) were significantly higher than that of the original genotype (382.0 mg/100 g). The highest total anthocyanin concentration was observed for the BSA078 genotype (467.9 mg/100 g). These results may be useful for identifying the optimal genotypes for breeding new cultivars with enhanced qualities and potential health benefits.