Geung-Joo Lee

Comparative Analysis of Miscanthus and Saccharum Reveals a Shared Whole-Genome Duplication but Different Evolutionary Fates.

This study reports a shared whole-genome duplication between two genera, Miscanthus and Saccharum. The genome duplication might have been a key event to shape different evolutionary fates of these two genera in terms of genomic/chromosomal structures and modes of reproduction. Multiple polyploidizations with divergent consequences in the grass subtribe Saccharinae provide a singular opportunity to study in situ adaptation of a genome to the duplicated state, heretofore known primarily from paleogenomics. We show that allopolyploidy in a common Miscanthus-Saccharum ancestor ∼3.8 to 4.6 million years ago closely coincides in time with their divergence from the Sorghum lineage. Subsequent Saccharum-specific autopolyploidy may have created pseudo-paralogous chromosome groups with random pairing within a group but infrequent pairing between groups. High chromosome number may reduce differentiation among Saccharum pseudo-paralogs by increasing opportunities for recombinations, with the lower chromosome numbers of Miscanthus favoring the return to disomic inheritance. The widespread tendency of plant chromosome numbers to recursively return to a narrow range following genome duplication appears to be occurring now in Saccharum spontaneum based on rich polymorphism for chromosome number among genotypes, with past reductions indicated by condensations of two ancestral chromosomes in Miscanthus (now n = 19) and perhaps as many as 10 in the Narenga-Sclerostachya clade (n = 15).

Sequencing of transcriptomes from two Miscanthus species reveals functional specificity in rhizomes, and clarifies evolutionary relationships.

BackgroundMiscanthus is a promising biomass crop for temperate regions. Despite the increasing interest in this plant, limited sequence information has constrained research into its biology, physiology, and breeding. The whole genome transcriptomes of M. sinensis and M. sacchariflorus presented in this study may provide good resources to understand functional compositions of two important Miscanthus genomes and their evolutionary relationships.ResultsFor M. sinensis, a total of 457,891 and 512,950 expressed sequence tags (ESTs) were produced from leaf and rhizome tissues, respectively, which were assembled into 12,166 contigs and 89,648 singletons for leaf, and 13,170 contigs and 112,138 singletons for rhizome. For M. sacchariflorus, a total of 288,806 and 267,952 ESTs from leaf and rhizome tissues, respectively, were assembled into 8,732 contigs and 66,881 singletons for leaf, and 8,104 contigs and 63,212 singletons for rhizome. Based on the distributions of synonymous nucleotide substitution (Ks), sorghum and Miscanthus diverged about 6.2 million years ago (MYA), Saccharum and Miscanthus diverged 4.6 MYA, and M. sinensis and M. sacchariflorus diverged 1.5 MYA. The pairwise alignment of predicted protein sequences from sorghum-Miscanthus and two Miscanthus species found a total of 43,770 and 35,818 nsSNPs, respectively. The impacts of striking mutations found by nsSNPs were much lower between sorghum and Miscanthus than those between the two Miscanthus species, perhaps as a consequence of the much higher level of gene duplication in Miscanthus and resulting ability to buffer essential functions against disturbance.ConclusionsThe ESTs generated in the present study represent a significant addition to Miscanthus functional genomics resources, permitting us to discover some candidate genes associated with enhanced biomass production. Ks distributions based on orthologous ESTs may serve as a guideline for future research into the evolution of Miscanthus species as well as its close relatives sorghum and Saccharum.

Botanical and germinating characteristics of Miscanthus species native to Korea

Rapid establishment by means of seed propagation is cost-effective for Miscanthus species, but the highest biomass-producing Miscanthus used worldwide predominantly is a sterile natural hybrid. Seeded variety and seed propagation will be desirable for extended cultivation of the species, but botanical features and optimal germination conditions of the potential Miscanthus species native to Korea have not been well compared and studied. The ploidy level of M. sinensis, M. sacchariflorus, and M. × giganteus used in this study were estimated to be diploidy, tetraploidy, and triploidy, which can be converted into nuclear DNA contents of 2.75 pg/C, 2.35 pg/C and 2.67 pg/C, respectively. M. sinensis rhizomes are compacted and less branching, while M. sacchariflorus has a broad creeping and thick stemmed rhizomes. Abaxial leaf surface of M. sinensis is densely pilose, but glabrous for M. sacchariflorus, while the leaf sheath of M. sacchariflorus is thickly haired, but hairless for M. sinensis. S pikelets of M. sinensis are awned, but awnless for M. sacchariflorus, which is one remarkable difference between them. When seed germination and seed loss by shattering are considered, the suggested harvest time is at least about 11th week after seed heading initiation. When seeds are not matured enough less than 10 weeks after seed heading, NaOCl treatment seems to damage seeds, resulting in lower seed germination. Compared to immature and less dried Miscanthus seeds, enhanced seed germination with 2% NaOCl, however, was evident for those matured seeds at most of the exposed temperatures ranging from 20 to 35. The greatest germination of 98 and 88% occurred at 30 for M. sinensis and M. sacchariflorus with 2% NaOCl scarification. From the effects of temperature on early growth, seedlings grown at 30 optimized shoot and root length, and fresh and dry weight for both Miscanthus species native to Korea.

Variation in the phenotypic features and transcripts of color mutants of chrysanthemum (Dendranthema grandiflorum) derived from gamma ray mutagenesis

We investigated the structural genes and their transcripts for anthocyanin synthesis inDendranthema grandiflorum ‘Argus’. Color variations in chrysanthemum mutants were obtained through gamm ray irradiation to regenerated plants from anin vitro. Normal florets were pinkish, but the mutants had white or purple ray florets and white, purple, or yellow-green disc florets. Irradiation modified both flower size and the number of ray florets. Compared with the control, levels of total anthocyanins in the mutants ranged from 4 times lower to 6 times higher for the disc florets. This disparity was even more evident, up to 14-fold greater, in the ray florets. Expression of the CHI, F3′H, F3′5′H, DFR, and LDOX genes varied among the mutants, but no dramatic changes were detected in CHS and F3H transcripts in either leaf or floret tissues. Sequence homology to known anthocyanin genes from other plant species was 61 to 84%, 62 to 74%, and 71 to 76% for CHI, F3′H, and LDOX, respectively. Our results support the proposal that such radiation-induced mutations in genes within the anthocyanin pathway are associated with variations in chrysanthemum flower color.

Analyses of genetic diversity and relationships in four Calanthe taxa native to Korea using AFLP markers

About 171 Calanthe species have been reported worldwide. However, only four species and two natural hybrids are known to grow naturally in South Korea. Classification studies of Calanthe orchids using molecular markers have been relatively limited. Therefore, the objectives of this study are to analyze the genetic diversity and relationships of four Calanthe taxa (101 accessions), including C. sieboldii (13 accessions), C. discolor (33), C. × bicolor (54), and C. aristulifera (1), native to South Korea and to identify the potential of each primer combination to discriminate each Calanthe accession using an AFLP technique. Thirty-two AFLP primer pairs produced a total of 2,764 bands, with an average of 86.4 bands per primer pair. Among all bands, 42.3% (1,354 bands), 51.6% (1,652), and 70.0% (2,240) were polymorphic in C. sieboldii, C. discolor, and C. × bicolor, respectively. Calanthe aristulifera was the most distant of the Calanthe taxa, and C. × bicolor, a natural hybrid of C. discolor × C. sieboldii, was more closely related to C. discolor (0.015 of genetic distance) than to C. sieboldii (0.022). Each Calanthe taxon with multiple accessions was divided into two or three groups. Comparisons of gene diversity, polymorphic information content, effective multiplex ratio, marker index, and resolving power proved that resolving power had the best discrimination potential for the 101 Calanthe accessions. These results will help to identify diverse accessions and to develop a breeding program of Calanthe orchids.

Identification of novel stress-induced microRNAs and their targets in Camelina sativa using computational approach

MicroRNAs (miRNAs) are a class of small non-coding, single-stranded RNA sequences that regulate gene expression at the post-transcriptional level and also reported to function in stress responses, but their role has not been studied in Camelina (Camelina sativa L.), an emerging oil crop. In this study, we predicted conserved as well as putative novel miRNAs from a Camelina drought stress cDNA library using comprehensive genomic approaches. Based on the sequence homology, we predicted 145 miRNAs, of which 61 were conserved, and 84 putative novel miRNAs were found to belong to 26 and 72 different miRNA families, respectively. In silico expression analysis indicated that 20 miRNAs were really expressed in Camelina genome, and several of them have tissue-specific expression character. We found that the 60 putative novel miRNA families target 117 genes. Most of the miRNA targets were predicted to genes including that regulate stress response, transcription factors, and fatty acid and lipid metabolism-related genes. Expression patterns of 6 randomly selected miRNAs under drought stress were validated by real-time quantitative polymerase chain reaction analysis. Coordinated expression changes between 6 randomly selected miRNAs and their target genes, suggested that the predicted miRNAs could be drought-responsive and that they would likely be directly involved in stress regulatory networks of Camelina. These results indicate that, in C. sativa, under drought stress, a large number of new miRNAs could be discovered, and the predicted stress-responsive miRNAs and their target transcripts will serve as valuable resources for future studies.

Targeted genome editing, an alternative tool for trait improvement in horticultural crops

Improving crops through plant breeding, an important approach for sustainable agriculture, has been utilized to increase the yield and quality of foods and other biomaterials for human use. Crops, including cereals, vegetables, ornamental flowers, fruits, and trees, have long been cultivated to produce high-quality products for human consumption. Conventional breeding technologies, such as natural cross-hybridization, mutation induction through physical or chemical mutagenesis, and modern transgenic tools are often used to enhance crop production. However, these breeding methods are sometimes laborious and complicated, especially when attempting to improve desired traits without inducing pleiotropic effects. Recently, targeted genome editing (TGE) technology using engineered nucleases, including meganucleases, zinc finger nucleases (ZFNs), transcription activator-like effector nucleases (TALENs), and clustered regularly interspaced short palindromic repeat (CRISPR) nucleases, has been used to improve the traits of economically important plants. TGE has emerged as a novel plant-breeding tool that represents an alternative approach to classical breeding, but with higher mutagenic efficiency. Here, we briefly describe the basic principles of TGE and the types of engineered nucleases utilized, along with their advantages and disadvantages. We also discuss their potential use to improve the traits of horticultural crops through genome engineering.

Genetic Diversity of Natural and Artificial Populations of Model Grass Brachypodium Species Evaluated by AFLP Markers

Brachypodium, a monocot grass with wide distribution in temperate areas, has been considered a new model plant for many grass species, such as turfgrass or miscanthus, due to its small genome size, self-pollination, rapid life cycle, higher seed yield, and small stature. The objectives of this study were to compare the genetic diversity of natural and artificial populations, and to determine the effects of γ-radiation on genetic variability when evaluated by amplified fragment length polymorphism (AFLP) markers. Two populations used in this study include 66 plant introductions (PI) from the U.S. Department of Agriculture (USDA; POP1), and 43 mutants derived from one of the plant introductions, Bd43 (PI 227011; POP2) treated with a γ-irradiation ranging from 100 to 600 Gy. The highest performance of the phenotypic traits was observed at 200 Gy for germination rate, 100 Gy for height, and 600 Gy for tiller number, implying that the LD50 for determining of optimum dosage depends on the physiological parameters measured. Based on AFLP analysis, POP2 showed higher polymorphism (79.4%), PIC (polymorphism information content; 0.162), and genetic diversity (0.195) than POP1 (59.4%, 0.113, and 0.130, respectively). Those three genetic parameters were evaluated for the mutants derived at various γ-radiation dosages and were found to be the highest when irradiated at 300 Gy. Artificial mutagenesis using γ-radiation improved genetic diversity compared to that of the natural population, and 300 Gy was a useful dosage to enlarge genetic variability in Brachypodium. Results suggest that a dosage maximizing the genetic diversity when evaluated with AFLP markers is an alternative index to LD50 for determining the optimum level for mutation induction.

Antioxidant and radical-scavenging activities of petal extracts of Camellia japonica ecotypes

Camellia japonica is the most well-known species of the genus Camellia, which is native to Korea, Japan, and China. In this study, the antioxidant and free radical-scavenging activities of petal extracts of Camellia ecotypes with different colors were evaluated. For total phenolic compounds, five ecotypes were grouped into low [(4.8 mg·g−1 of dry weight (DW) GAE (gallic acid equivalent)], medium (6.0–6.2 mg·g−1) and high (19.6 mg·g−1) groups for white, red, and pink colors, respectively. The DPPH radical scavenging activity of the petal extracts, presented as a half maximal inhibitory concentration (IC50), was the greatest (3.8 μg·mL−1) in the pink ecotype and least (43.1 μg·mL−1) in the white ecotype as compared to ascorbic acid (13.6 μg·mL−1). The results demonstrated that the efficient DPPH radical scavenging activity of the pink ecotype was partly due to higher contents of phenolic compounds. Activities of two antioxidant enzymes, catalase and peroxidase, varied among the ecotypes, indicating the presence of ecotype-specific detoxifying processes. Camellia petals had higher amounts of antioxidants as well as excellent reactive oxygen species (ROS) scavenging activities that were dependent on the petal color.

Characterization of Salt Tolerant Rice Mutant Lines Derived from Azetidine-2-Carboxylic Acid Resistant Cell Lines Induced by Gamma Ray Irradiation

To develop rice (Oryza sativa L.) cultivars to be planted on salt-affected sites, cell lines with enhanced proline content and resistance to growth inhibition by Azetidine-2-carboxylic acid (AZCA), a proline analogue, were screened out among calli irradiated with gamma ray of 50, 70, 90, and 120 Gy. The calli had been derived from embryo culture of the cultivar Donganbyeo. Selected AZCA resistant lines that had high proline accumulation were used as sources for selection of NaCl resistant lines. To determine an optimum concentration for selection of NaCl resistant lines, Donganbyeo seeds were initially cultured on the media containing various NaCl concentrations (0 to 2.5%) for 40 days, and 1.5% NaCl concentration was determined as the optimum concentration. One hundred sixteen salt-tolerant (ST) lines were selected from bulked 20,000 seeds of the AZCA resistant seeds in the medium containing 1.5% NaCl. The putative 33 lines ( generation) considered with salt-tolerance were further analyzed for salt tolerance, amino acid and ion contents, and expression patterns of the salt tolerance-related genes. Out of the 33 lines, 7 lines were confirmed to have superior salt tolerance. Based on growth comparison of the entries, the selected mutant lines exhibited greater shoot length with average 1.5 times, root length with 1.3 times, root numbers with 1.1 times, and fresh weight with 1.5 times than control. Proline contents were increased maximum 20%, 100% and 20% in the leaf, seed and callus, respectively, of the selected lines. Compared to control, amino acid contents of the mutants were 24 to 29%, 49 to 143%, 32 to 60% higher in the leaf, seed and callus, respectively. The ratio of for most of the ST-lines were lower than that of control, ranging from 1.0 to 3.8 for the leaf and 11.5 to 28.5 for the root, while the control had 3.5 and 32.9 in the leaf and root, respectively. The transcription patterns for the P5CS and NHXI genes observed by RT-PCR analysis indicated that these genes were actively expressed under salt stress. The selected mutants will be useful for the development of rice cultivar resistant to salt stress.