Genome-Wide Association Study of Phenolic Content and Antioxidant Properties in Eggplant Germplasm.

The phenolic compounds in eggplant offer potential natural antioxidants for improved health. A large number of samples were examined in order to find eggplant germplasm with a high potential for health promotion. A genome-wide association study (GWAS) was conducted to identify single nucleotide polymorphisms (SNPs) associated with variations in total phenolic content (TPC) and antioxidant activity in eggplants, including ABTS (2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid)) scavenging activity and ferric reducing antioxidant power (FRAP). TPC values varied from 14.19 to 842.90 mg gallic acid equivalent (GAE)/100 g of dry weight of eggplant fruit powder. TPC showed a strong positive correlation with both FRAP and ABTS (r = 0.89 *** and 0.77 ***, respectively). The GWAS identified 20 SNPs that were significantly associated out of 29,183 SNPs. Out of the 20 significant SNPs, 11 showed associations with TPC, 4 with ABTS activity, and 5 with FRAP. Among the SNPs associated with TPC, one SNP was found on each of Chromosomes 3, 4, 7, and 12. In contrast, Chromosome 5 comprised two SNPs associated to TPC. Furthermore, the gene encoding IRX12 laccase-4 on Chromosome 10 was found to contain five SNPs associated with TPC. Four significantly linked SNPs on Chromosomes 1 (1 SNP), 4 (2 SNPs), and 10 (1 SNP) were found to be related to ABTS activity. The identified SNPs will be further examined as markers for selecting desirable eggplant varieties and exploring the links between candidate genes, phenolic content, and antioxidant activity. The findings of this study could assist in further study and the development of eggplants with improved health advantages through targeted breeding.

Genetic Stability Assessment of Six Cryopreserved Strawberry (Fragaria × ananassa Duch.) Accessions by Phenotypic and Molecular Studies.

For the long-term preservation of genetic resources, cryopreservation techniques have been developed for strawberry germplasm, mainly using in vitro-grown shoot tips. In this study, genetic stability was tested under greenhouse conditions for six strawberry accessions (IT232511, PHS0132, IT245810, IT245830, IT245852, and IT245860) derived from the following procedures: (1) conventional propagation (GH: greenhouse maintained); (2) in vitro propagation (TC: tissue culture); (3) pretreatment before cryopreservation (-LN: non-liquid nitrogen exposure); and (4) cryopreservation (+LN: liquid nitrogen exposure). To test the performance of phenotypic traits, we measured six vegetative and five fruit traits. There were no distinct differences in most of the characteristics, but a few traits, such as sugar content and pH of fruits in three accessions, showed higher values in +LN compared to GH. However, the differences disappeared in the first runner generation. To test genetic variations, a total of 102 bands were generated by twelve inter simple sequence repeat (ISSR) primers. A few polymorphic bands were found only in plants derived from TC of IT245860, which was not cryopreserved. The sequencing analysis of four polymorphic bands produced by ISSR_15 showed that none of these sequences matched the characterized genes in NCBI. Phenotypic abnormality was not observed across all plants. This study indicates that cryopreserved plants of the six strawberry accessions are phenotypically and genetically stable. Therefore, the results of this study can help to implement cryobanking of strawberry germplasm.

Genome-Wide Association Study for Agro-Morphological Traits in Eggplant Core Collection.

Eggplant is one of the most economically and nutritionally important vegetables worldwide. The study of the association of phenotypic traits with genetic factors is vital for the rapid and efficient identification and selection of eggplant genetic resources for breeding purposes with desired traits. The eggplant resources (587) collected from different countries, including Korea, were used for establishing the core collection. A total of 288 accessions were selected from 587 Solanum accessions based on 52 single nucleotide polymorphisms (SNPs) markers together with 17 morphological traits. This core collection was further used to analyze the genetic associations of eggplant morphological variations. A large variation was found among the evaluated eggplant accessions for some agro-morphological traits. Stem prickles and leaf prickles showed a significant positive correlation (r = 0.83***), followed by days to flowering and days to maturity (r = 0.64***). A total of 114,981 SNPs were filtered and used for phylogenetic tree analysis, population structure analysis, and genome-wide association study (GWAS). Among the agro-morphological traits, significantly associated SNPs were found for six traits. A total of 377 significantly associated SNPs with six agro-morphological traits were identified. These six traits and the number of SNPs were: days to maturity (51), flower size (121), fruit width (20), harvest fruit color (42), leaf prickles (38), and stem prickles (105). The largest fraction of significant SNPs (11.94%) was obtained on chromosome Ch01, followed by Ch07 and Ch06 with 11.67% and 10.08%, respectively. This study will help to develop markers linked to the most important agro-morphological traits of eggplant genetic resources and support the selection of desirable traits for eggplant breeding programs.

Interspecific variation of seed morphological and micro-morphological traits in the genus Vicia (Fabaceae).

Seed macro- and micro-morphology were analyzed to evaluate their capacity to discriminate species in the genus Vicia (Fabaceae). To assess the interspecific variation of the taxa in the genus Vicia, 41 accessions were obtained from the USDA-ARS germplasm collection in the USA and 19 accessions were collected from Korea. Seed morphological characteristics such as shape, color, mottling, finish, length, width, diameter, hilum shape, hilum color, hilum length, and lens distance from the hilum were examined under a stereomicroscope. Testa texture characteristics such as testa pattern, papillae type, density, height, ribbing, surface deposits, and peaks topped with wax were examined under scanning electron microscopy. Various gross-morphological traits of seeds of Vicia species have been analyzed and compared. The present study revealed significant variation in testa traits. Testa were papillose and papillose with mounds, the latter being observed only in Vicia lathyroides. The present study revealed 20 key traits that could be used to diagnose Vicia species and classify them.

Genotyping-by-Sequencing Derived Single Nucleotide Polymorphisms Provide the First Well-Resolved Phylogeny for the Genus Triticum (Poaceae).

Wheat (Triticum spp.) has been an important staple food crop for mankind since the beginning of agriculture. The genus Triticum L. is composed of diploid, tetraploid, and hexaploid species, majority of which have not yet been discriminated clearly, and hence their phylogeny and classification remain unresolved. Genotyping-by-sequencing (GBS) is an easy and affordable method that allows us to generate genome-wide single nucleotide polymorphism (SNP) markers. In this study, we used GBS to obtain SNPs covering all seven chromosomes from 283 accessions of Triticum-related genera. After filtering low-quality and redundant SNPs based on haplotype information, the GBS assay provided 14,188 high-quality SNPs that were distributed across the A (71%), B (26%), and D (2.4%) genomes. Cluster analysis and discriminant analysis of principal components (DAPC) allowed us to distinguish six distinct groups that matched well with Triticum species complexity. We constructed a Bayesian phylogenetic tree using 14,188 SNPs, in which 17 Triticum species and subspecies were discriminated. Dendrogram analysis revealed that the polyploid wheat species could be divided into groups according to the presence of A, B, D, and G genomes with strong nodal support and provided new insight into the evolution of spelt wheat. A total of 2,692 species-specific SNPs were identified to discriminate the common (T. aestivum) and durum (T. turgidum) wheat cultivar and landraces. In principal component analysis grouping, the two wheat species formed individual clusters and the SNPs were able to distinguish up to nine groups of 10 subspecies. This study demonstrated that GBS-derived SNPs could be used efficiently in genebank management to classify Triticum species and subspecies that are very difficult to distinguish by their morphological characters.

Phytochemicals and Antioxidant Activity of Korean Black Soybean (Glycine max L.) Landraces.

Black soybean (Glycine max L.) has been used as a traditional medicine because its seed coat contains various natural phenolic compounds such as anthocyanins. The objective of this study was to reveal the genetic variation in the agricultural traits, phytochemicals, and antioxidant activity of 172 Korean black soybean landraces (KBSLs) and establish a relationship among them. The evaluation of three agricultural traits (days to 50% flowering, maturity, and 100-seed weight), six phytochemicals (delphinidin-3-glucoside, cyaniding-3-glucoside, petunidin-3-glucoside, daidzin, glycitin, and genestin), and four antioxidant activities (2,2-diphenyl-1-picrylhydrazyl (DPPH), 2,2'-azino-bis-(3-ethylbenzothiazoline-6-sulfonic acid)(ABTS), ferric-reducing antioxidant power (FRAP), and the total polyphenol content (TPC) of 172 KBSLs were analyzed in 2012 and 2015. The agricultural traits, phytochemicals, and antioxidant activities of the 172 KBSLs showed wide variation among the accessions and years. In correlation analysis, the agricultural traits and phytochemicals showed positive and negative correlations with phytochemicals and antioxidant activity, respectively. The principal component analyses result indicated that phytochemicals accounted for most of the variability in the KBSLs. In clustering analysis, the 172 KBSLs were classified into four clusters. These results could lead to expanding the knowledge of the agricultural traits, phytochemicals, and antioxidant activity of the KBSLs, which are valuable materials for the development of new soybean varieties.

Genetic Diversity Assessed by Genotyping by Sequencing (GBS) in Watermelon Germplasm.

Watermelon is an economically important vegetable fruit worldwide. The objective of this study was to conduct a genetic diversity of 68 watermelon accessions using single nucleotide polymorphisms (SNPs). Genotyping by sequencing (GBS) was used to discover SNPs and assess genetic diversity and population structure using STRUCTURE and discriminant analysis of principal components (DAPC) in watermelon accessions. Two groups of watermelons were used: 1) highly utilized 41 watermelon accessions at the National Agrobiodiversity Center (NAC) at the Rural Development Administration in South Korea; and 2) 27 Korean commercial watermelons. Results revealed the presence of four clusters within the populations differentiated principally based on seed companies. In addition, there was higher genetic differentiation among commercial watermelons of each company. It is hypothesized that the results obtained from this study would contribute towards the expansion of this crop as well as providing data about genetic diversity, which would be useful for the preservation of genetic resources or for future breeding programs.

The complete chloroplast genome sequence of wild oat, Avena sterilis L. (Poaceae) and its phylogeny.

Wild oat, Avena sterilis L. is a stout broad-leaved annual grass resembling cultivated oats in general appearance. In this study, we sequenced the complete chloroplast (cp) genome sequence of A. sterilis for the first time to investigate their phylogenetic relationship in the family Poaceae. The complete cp genome sequence is 135,887 bp in length with 38.5% overall GC content and exhibits a typical quadripartite structure comprising one pair of inverted repeats (21,603 bp) separated by a small single-copy region (12,575 bp) and a large single-copy region (80,106). The cp genome encodes 111 unique genes, 76 of which are protein-coding genes, four rRNA genes, 30 tRNA genes, and 18 duplicated genes in the inverted repeat region. The phylogenetic analysis indicated A. sterilis closely clustered with the cultivated oat, A. sativa L.

The complete chloroplast genome sequences of little millet (Panicum sumatrense Roth ex Roem. and Schult.) (Poaceae).

Little millet, Panicum sumatrense Roth ex Roem. & Schult., is an important cultivated species under the tribe Paniceae, sub-family Panicoideae and family Poaceae. In this study, for the first time we sequenced the complete chloroplast (cp) genome of P. sumatrense to investigate their phylogenetic relationship in the family Poaceae. The complete cp genome sequence of P. sumatrense is 139,384 bp in length with 38.6% overall GC content and exhibits a typical quadripartite structure comprising one pair of inverted repeats (22,723 bp) separated by a small single-copy region (12,583 bp) and a large single-copy region (81,355 bp). The P. sumatrense cp genome encodes 125 unique genes, which include 91 protein-coding genes, 4 rRNA genes, 30 tRNA genes, and 20 genes were duplicated in the inverted repeat region. This newly determined cp genome (P. sumatrense) could be valuable information for the breeding programs of this cereal crops in the family Poaceae.

New Genetic Loci Associated with Preharvest Sprouting and Its Evaluation Based on the Model Equation in Rice.

Preharvest sprouting (PHS) in rice panicles is an important quantitative trait that causes both yield losses and the deterioration of grain quality under unpredictable moisture conditions at the ripening stage. However, the molecular mechanism underlying PHS has not yet been elucidated. Here, we explored the genetic loci associated with PHS in rice and formulated a model regression equation for rapid screening for use in breeding programs. After re-sequencing 21 representative accessions for PHS and performing enrichment analysis, we found that approximately 20,000 SNPs revealed distinct allelic distributions between PHS resistant and susceptible accessions. Of these, 39 candidate SNP loci were selected, including previously reported QTLs. We analyzed the genotypes of 144 rice accessions to determine the association between PHS and the 39 candidate SNP loci, 10 of which were identified as significantly affecting PHS based on allele type. Based on the allele types of the SNP loci, we constructed a regression equation for evaluating PHS, accounting for an R2 value of 0.401 in japonica rice. We validated this equation using additional accessions, which exhibited a significant R2 value of 0.430 between the predicted values and actual measurements. The newly detected SNP loci and the model equation could facilitate marker-assisted selection to predict PHS in rice germplasm and breeding lines.

The complete chloroplast genome of Capsicum frutescens (Solanaceae).

PREMISE OF THE STUDY: We report the complete sequence of the chloroplast genome of Capsicum frutescens (Solanaceae), a species of chili pepper. METHODS AND RESULTS: Using an Illumina platform, we sequenced the chloroplast genome of C. frutescens. The total length of the genome is 156,817 bp, and the overall GC content is 37.7%. A pair of 25,792-bp inverted repeats is separated by small (17,853 bp) and large (87,380 bp) single-copy regions. The C. frutescens chloroplast genome encodes 132 unique genes, including 87 protein-coding genes, 37 transfer RNA (tRNA) genes, and eight ribosomal RNA (rRNA) genes. Of these, seven genes are duplicated in the inverted repeats and 12 genes contain one or two introns. Comparative analysis with the reference chloroplast genome revealed 125 simple sequence repeat motifs and 34 variants, mostly located in the noncoding regions. CONCLUSIONS: The complete chloroplast genome sequence of C. frutescens reported here is a valuable genetic resource for Capsicum species.

Transcriptome Analysis of Two Vicia sativa Subspecies: Mining Molecular Markers to Enhance Genomic Resources for Vetch Improvement.

The vetch (Vicia sativa) is one of the most important annual forage legumes globally due to its multiple uses and high nutritional content. Despite these agronomical benefits, many drawbacks, including cyano-alanine toxin, has reduced the agronomic value of vetch varieties. Here, we used 454 technology to sequence the two V. sativa subspecies (ssp. sativa and ssp. nigra) to enrich functional information and genetic marker resources for the vetch research community. A total of 86,532 and 47,103 reads produced 35,202 and 18,808 unigenes with average lengths of 735 and 601 bp for V. sativa sativa and V. sativa nigra, respectively. Gene Ontology annotations and the cluster of orthologous gene classes were used to annotate the function of the Vicia transcriptomes. The Vicia transcriptome sequences were then mined for simple sequence repeat (SSR) and single nucleotide polymorphism (SNP) markers. About 13% and 3% of the Vicia unigenes contained the putative SSR and SNP sequences, respectively. Among those SSRs, 100 were chosen for the validation and the polymorphism test using the Vicia germplasm set. Thus, our approach takes advantage of the utility of transcriptomic data to expedite a vetch breeding program.

The complete chloroplast genome of Capsicum annuum var. glabriusculum using Illumina sequencing.

Chloroplast (cp) genome sequences provide a valuable source for DNA barcoding. Molecular phylogenetic studies have concentrated on DNA sequencing of conserved gene loci. However, this approach is time consuming and more difficult to implement when gene organization differs among species. Here we report the complete re-sequencing of the cp genome of Capsicum pepper (Capsicum annuum var. glabriusculum) using the Illumina platform. The total length of the cp genome is 156,817 bp with a 37.7% overall GC content. A pair of inverted repeats (IRs) of 50,284 bp were separated by a small single copy (SSC; 18,948 bp) and a large single copy (LSC; 87,446 bp). The number of cp genes in C. annuum var. glabriusculum is the same as that in other Capsicum species. Variations in the lengths of LSC; SSC and IR regions were the main contributors to the size variation in the cp genome of this species. A total of 125 simple sequence repeat (SSR) and 48 insertions or deletions variants were found by sequence alignment of Capsicum cp genome. These findings provide a foundation for further investigation of cp genome evolution in Capsicum and other higher plants.

Novel microsatellite markers acquired from Rubus coreanus Miq. and cross-amplification in other Rubus species.

The Rubus genus consists of more than 600 species that are distributed globally. Only a few Rubus species, including raspberries and blueberries, have been domesticated. Genetic diversity within and between Rubus species is an important resource for breeding programs. We developed genomic microsatellite markers using an SSR-enriched R. coreanus library to study the diversity of the Rubus species. Microsatellite motifs were discovered in 546 of 646 unique clones, and a dinucleotide repeat was the most frequent (75.3%) type of repeat. From 97 microsatellite loci with reproducible amplicons, we acquired 29 polymorphic microsatellite markers in the Rubus coreanus collection. The transferability values ranged from 59.8% to 84% across six Rubus species, and Rubus parvifolius had the highest transferability value (84%). The average number of alleles and the polymorphism information content were 5.7 and 0.541, respectively, in the R. coreanus collection. The diversity index of R. coreanus was similar to the values reported for other Rubus species. A phylogenetic dendrogram based on SSR profiles revealed that seven Rubus species could be allocated to three groups, and that R. coreanus was genetically close to Rubus crataegifolius (mountain berry). These new microsatellite markers might prove useful in studies of the genetic diversity, population structure, and evolutionary relationships among Rubus species.

Cross-amplification of Vicia sativa subsp. sativa microsatellites across 22 other Vicia species.

The temperate and herbaceous genus Vicia L. is a member of the legume tribe Fabeae of the subfamily Papilionoideae. The genus Vicia comprises 166 annual or perennial species distributed mainly in Europe, Asia, and North America, but also extending to the temperate regions of South America and tropical Africa. The use of simple sequence repeat (SSR) markers for Vicia species has not been investigated as extensively as for other crop species. In this study, we assessed the potential for cross-species amplification of cDNA microsatellite markers developed from common vetch (Vicia sativa subsp. sativa). For cross-species amplification of the SSRs, amplification was carried out with genomic DNA isolated from two to eight accessions of 22 different Vicia species. For individual species or subspecies, the transferability rates ranged from 33% for V. ervilia to 82% for V. sativa subsp. nigra with an average rate of 52.0%. Because the rate of successful SSR marker amplification generally correlates with genetic distance, these SSR markers are potentially useful for analyzing genetic relationships between or within Vicia species.

Development of a core set from a large rice collection using a modified heuristic algorithm to retain maximum diversity.

A new heuristic approach was undertaken for the establishment of a core set for the diversity research of rice. As a result, 107 entries were selected from the 10 368 characterized accessions. The core set derived using this new approach provided a good representation of the characterized accessions present in the entire collection. No significant differences for the mean, range, standard deviation and coefficient of variation of each trait were observed between the core and existing collections. We also compared the diversity of core sets established using this Heuristic Core Collection (HCC) approach with those of core sets established using the conventional clustering methods. This modified heuristic algorithm can also be used to select genotype data with allelic richness and reduced redundancy, and to facilitate management and use of large collections of plant genetic resources in a more efficient way.