Development and Validation of SNP and InDel Markers for Pod-Shattering Tolerance in Soybean.

Pod-shattering causes a significant yield loss in many soybean cultivars. Shattering-tolerant cultivars provide the most effective approach to minimizing this loss. We developed molecular markers for pod-shattering and validated them in soybeans with diverse genetic backgrounds. The genes Glyma.16g141200, Glyma.16g141500, and Glyma.16g076600, identified in our previous study by quantitative trait locus (QTL) mapping and whole-genome resequencing, were selected for marker development. The whole-genome resequencing of three parental lines (one shattering-tolerant and two shattering-susceptible) identified single nucleotide polymorphism (SNP) and/or insertion/deletion (InDel) regions within or near the selected genes. Two SNPs and one InDel were converted to Kompetitive Allele-Specific PCR (KASP) and InDel markers, respectively. The accuracy of the markers was examined in the two recombinant inbred line populations used for the QTL mapping, as well as the 120 varieties and elite lines, through allelic discrimination and phenotyping by the oven-drying method. Both types of markers successfully discriminated the pod shattering-tolerant and shattering-susceptible genotypes. The prediction accuracy, which was as high as 90.9% for the RILs and was 100% for the varieties and elite lines, also supported the accuracy and usefulness of these markers. Thus, the markers can be used effectively for genetic and genomic studies and the marker-assisted selection for pod-shattering tolerance in soybean.

Protein, Amino Acid, Oil, Fatty Acid, Sugar, Anthocyanin, Isoflavone, Lutein, and Antioxidant Variations in Colored Seed-Coated Soybeans.

Different physiological and genetic studies show that the variations in the accumulation of pigment-stimulating metabolites result in color differences in soybean seed coats. The objective of this study was to analyze the nutrient contents and antioxidant potential in black, brown, and green seed-coated soybeans. Significant variations in protein (38.9-43.3%), oil (13.9-20.4%), total sugar (63.5-97.0 mg/g seed), total anthocyanin (3826.0-21,856.0 µg/g seed coat), total isoflavone (709.5-3394.3 µg/g seed), lutein (1.9-14.8 µg/g), total polyphenol (123.0-385.8 mg gallic acid/100 g seed), total flavonoid (22.1-208.5 mg catechin/100 g seed), 2,2'-azino-bis(3-ethylbenzthiazoline-6-sulphonic acid (ABTS; 275.0-818.8 mg Trolox/100 g seed), and 2,2-diphenyl-1-picrylhydrazyl (DPPH; 96.3-579.7 mg Trolox/100 g seed) were found among the soybean genotypes. Ilpumgeomjeong2 contained the lowest protein but the highest oil and total sugar. The lowest oil-containing Wonheug had the highest protein content. Socheong2 was rich in all four variables of antioxidants. Anthocyanins were detected only in black soybeans but not in brown and green soybeans. The variation in isoflavone content was up to 5-fold among the soybean genotypes. This study could be a valuable resource for the selection and improvement of soybean because an understanding of the nutrient content and antioxidant potentials is useful to develop effective strategies for improving the economic traits; for example, the major emphasis of soybean breeding for fatty acids is to enhance the oleic and linoleic acid contents and to decrease linolenic acid content.

Quantitative Trait Locus Mapping for Drought Tolerance in Soybean Recombinant Inbred Line Population.

Improving drought stress tolerance of soybean could be an effective way to minimize the yield reduction in the drought prevailing regions. Identification of drought tolerance-related quantitative trait loci (QTLs) is useful to facilitate the development of stress-tolerant varieties. This study aimed to identify the QTLs for drought tolerance in soybean using a recombinant inbred line (RIL) population developed from the cross between a drought-tolerant 'PI416937' and a susceptible 'Cheonsang' cultivar. Phenotyping was done with a weighted drought coefficient derived from the vegetative and reproductive traits. The genetic map was constructed using 2648 polymorphic SNP markers that distributed on 20 chromosomes with a mean genetic distance of 1.36 cM between markers. A total of 10 QTLs with 3.52-4.7 logarithm of odds value accounting for up to 12.9% phenotypic variance were identified on seven chromosomes. Five chromosomes-2, 7, 10, 14, and 20-contained one QTL each, and chromosomes 1 and 19 harbored two and three QTLs, respectively. The chromosomal locations of seven QTLs overlapped or located close to the related QTLs and/or potential candidate genes reported earlier. The QTLs and closely linked markers could be utilized in maker-assisted selection to accelerate the breeding for drought tolerance in soybean.

QTL Mapping and Candidate Gene Analysis for Pod Shattering Tolerance in Soybean (Glycine max).

Pod shattering is an important reproductive process in many wild species. However, pod shattering at the maturing stage can result in severe yield loss. The objectives of this study were to discover quantitative trait loci (QTLs) for pod shattering using two recombinant inbred line (RIL) populations derived from an elite cultivar having pod shattering tolerance, namely "Daewonkong", and to predict novel candidate QTL/genes involved in pod shattering based on their allele patterns. We found several QTLs with more than 10% phenotypic variance explained (PVE) on seven different chromosomes and found a novel candidate QTL on chromosome 16 (qPS-DS16-1) from the allele patterns in the QTL region. Out of the 41 annotated genes in the QTL region, six were found to contain SNP (single-nucleotide polymorphism)/indel variations in the coding sequence of the parents compared to the soybean reference genome. Among the six potential candidate genes, Glyma.16g076600, one of the genes with known function, showed a highly differential expression levels between the tolerant and susceptible parents in the growth stages R3 to R6. Further, Glyma.16g076600 is a homolog of AT4G19230 in Arabidopsis, whose function is related to abscisic acid catabolism. The results provide useful information to understand the genetic mechanism of pod shattering and could be used for improving the efficiency of marker-assisted selection for developing varieties of soybeans tolerant to pod shattering.

Fruit Morphology, Citrulline, and Arginine Levels in Diverse Watermelon (Citrullus lanatus) Germplasm Collections.

Watermelon (Citrullus lanatus) is a non-seasonal, economically important, cucurbit cultivated throughout the world, with Asia as a continent contributing the most. As part of the effort to diversify watermelon genetic resources in the already cultivated group, this study was devoted to providing baseline data on morphological quality traits and health-beneficial phytonutrients of watermelon germplasm collections, thereby promoting watermelon research and cultivation programs. To this end, we reported morphological traits, citrulline, and arginine levels of watermelon genetic resources obtained from the gene bank of Agrobiodiversity Center, Republic of Korea, and discussed the relationships between each. Diverse characteristics were observed among many of the traits, but most of the genetic resources (>90%) were either red or pink-fleshed. Korean originated fruits contained intermediate levels of soluble solid content (SSC) while the USA, Russian, Tajikistan, Turkmenistan, Taiwan, and Uruguay originated fruits had generally the highest levels of soluble solids. The citrulline and arginine contents determined using the High Performance Liquid Chromatography (HPLC) method ranged from 6.9 to 52.1 mg/g (average, 27.3 mg/g) and 1.8 to 21.3 mg/g (average, 9.8 mg/g), respectively. The citrulline content determined using the Citrulline Assay Kit ranged from 6.5 to 42.8 mg/g (average, 27.0 mg/g). Resources with high citrulline and arginine levels contained low SSC, whereas red- and pink-colored flesh samples had less citrulline compared to yellow and orange.

Identification and quantification of selected metabolites in differently pigmented leaves of lettuce (Lactuca sativa L.) cultivars harvested at mature and bolting stages.

BACKGROUND: Identification and screening of cultivars rich in bioactive phytoconstituents can be potentially useful to make nutrient-dense dishes and in medicinal formulations. In this study, we have identified, characterized and quantified caffeoylquinic acids, dicaffeoylquinic acid, dicaffeoyltartaric acid, kaempferol conjugates, quercetin malonylglucoside, sesquiterpene lactones, and cyanidin in 22 lettuce cultivars at mature and bolting stages using UPLC-PDA-Q-TOF-HDMS, UPLC, and HPLC. RESULTS: The composition and contents of the studied metabolites and antioxidant activity varied significantly and depend on leaf color, cultivar type and stage of maturity. The main phenolic acid components of lettuce were quinic and tartaric acid derivatives, whereas kaempferol derivatives were the dominant flavonoids. The sum of the content of phenolic acids ranged from 18.3 to 54.6 mg/100 g DW and 15.5 to 54.6 mg/100 g DW, whereas the sum of the contents of flavonoids ranged from 9.2 to 25.9 mg/100 g DW and 14.9 to 83.0 mg/100 g DW in mature and bolting stage cultivars, respectively. The content of cyanidin, lactucin, lactucopicrin, and ABTS radical antioxidant activity were in the range of 0.3 to 9.7 (mature stage) and 0.5 to 10.2 mg/g DW (bolting stage), 1.8 to 41.9 (mature stage) and 9.7 to 213.0 (bolting stage) µg/g DW, 9.9 to 344.8 (mature stage) and 169.2 to 3888.2 (bolting stage) µg/g DW, and 12.1 to 29.0 (mature stage) and 15.7 to 30.3 (bolting stage) mg TE/g DW, respectively. The principal component analysis (PCA) showed that the green and red pigmented lettuce cultivars were grouped to the negative and positive sides of PC1, respectively, while the green/red pigmented cultivars were distributed throughout the four quadrants of the PCA plots with no prominent grouping. The loading plot showed that phenolic acids, flavonoids, and cyanidin are the most potent contributors to the radical scavenging activity of lettuce extracts. CONCLUSIONS: Lettuce at the bolting stage accumulate relatively high amount of sesquiterpene lactones (SLs), quercetin malonylglucoside (QMG), methylkaempferol glucuronide (MKGR), kaempferol malonylglucoside (KMG), and 3-O-caffeoylquinic acid (3-CQA) compared to the mature stage. Higher amount of phytoconstituents were found to be accumulated in the red pigmented lettuce leaves compared to the green lettuce leaves. In addition, the contents of most of the metabolites in lettuce seem to increase with age of the leaves. The presence of the two bitter SLs, lactucin and lactucopicrin, in significantly high amount in lettuce leaves at bolting stage could diminish consumer acceptance. However, alternatively, these leaves could be utilized by nutraceutical companies working to recover these compounds.

Evaluation of Resistance to Ralstonia solanacearum in Tomato Genetic Resources at Seedling Stage.

Bacterial wilt of tomatoes caused by Ralstonia solanacearum is a devastating disease that limits the production of tomato in Korea. The best way to control this disease is using genetically resistant tomato plant. The resistance degree to R. solanacearum was evaluated for 285 tomato accessions conserved in the National Agrobiodiversity Center of Rural Development Administration. These accessions of tomato were originated from 23 countries. Disease severity of tomato accessions was investigated from 7 days to 14 days at an interval of 7 days after inoculation of R. solanacearum under greenhouse conditions. A total of 279 accessions of tomato germplasm were susceptible to R. solanacearum, resulting in wilt and death in 70 to 90% of these plants. Two tomato accessions were moderately resistant to R. solanacearum. Only four accessions showed high resistance against R. solanacearum. No distinct symptom of bacterial wilt appeared on the resistant tomato germplasms for up to 14 days after inoculation of R. solanacearum. Microscopy of resistant tomato stems infected with R. solanacearum revealed limited bacterial spread with thickening of pit membrane and gum production. Therefore, these four resistant tomato germplasms could be used in tomato breeding program against bacterial wilt.

Genetic assessment of safflower (Carthamus tinctorius L.) collection with microsatellite markers acquired via pyrosequencing method.

A genetic evaluation of safflower germplasm collections derived from different geographical regions and countries will provide useful information for sustainable conservation and the utilization of genetic diversity. However, the molecular marker information is limited for evaluation of genetic diversity of safflower germplasm. In this study, we acquired 509 putative genomic SSR markers for sufficient genome coverage using next-generation sequencing methods and characterized thirty polymorphic SSRs in safflower collection composed of 100 diverse accessions. The average allele number and expected heterozygosity were 2.8 and 0.386, respectively. Analysis of population structure and phylogeny based on thirty SSR profiles revealed genetic admixture between geographical regions contrary to genetic clustering. However, the accessions from Korea were genetically conserved in distinctive groups in contrast to other safflower gene pool. In conclusion, these new genomic SSRs will facilitate valuable studies to clarify genetic relationships as well as conduct population structure analyses, genetic map construction and association analysis for safflower.

Incidence, Molecular Characteristics and Pathogenicity of Gibberella fujikuroi Species Complex Associated with Rice Seeds from Asian Countries.

Gibberella fujikuroi species complex (GFSC) was isolated from rice (Oryza sativa L.) seed samples from ten Asian countries and investigated for incidence of GFSC, molecular characteristics, and pathogenicity. Regardless of geographic origin, GFSC was detected with incidences ranging from 3% to 80%. Four species, Fusarium fujikuroi, F. concentricum, F. proliferatum, and F. verticillioides, were found to show an association with rice seeds, with F. fujikuroi being the predominant species. In phylogenetic analyses of DNA sequences, no relationship was found between species, isolates, and geographic sources of samples. Unidentified fragments of the ß-tubulin gene were observed in ten isolates of F. fujikuroi and F. verticillioides. With the exception of three isolates of F. fujikuroi, F. fujikuroi, F. proliferatum, and F. verticillioides were found to have FUM1 (the fumonisin biosynthetic gene); however, FUM1 was not found in isolates of F. concentricum. Results of pathogenicity testing showed that all isolates caused reduced germination of rice seed. In addition, F. fujikuroi and F. concentricum caused typical symptoms of bakanae, leaf elongation and chlorosis, whereas F. proliferatum and F. verticillioides only caused stunting of seedlings. These findings provide insight into the characteristics of GFSC associated with rice seeds and might be helpful in development of strategies for management of bakanae.

Molecular authentication of 21 Korean artemisia species (Compositae) by polymerase chain reaction-restriction fragment length polymorphism based on trnL-F region of chloroplast DNA.

The present study describes the molecular authentication of 21 Korean Artemisia species using PCR-RFLP (polymerase chain reaction-restriction fragment length polymorphism) technique based on the trnL-F sequences in chloroplast DNA. Five different banding patterns were generated from 21 Artemisia species using HinfI restriction enzyme. A. apiacea, A. keiskeana and A. sieversiana have specific banding patterns. The remaining 18 species had shared two banding patterns. Phylogenetic analysis based on trnL-F sequence variations showed results similar to PCR-RFLP banding patterns. It suggested that the trnL-F region does not have sufficient variations to identify the 21 Artemisia species. However, the specific banding patterns for A. apiacea, A. keiskeana and A. sieversiana can be utilized as a DNA marker for discriminating them from other Artemisia species. These markers will be also useful for developing A. apiacea, A. keiskeana and A. sieversiana into new medicine and food based on their efficacy.