Background : Codonopsis lanceolata is used as a natural medicine or vegetables. It originates in East Asia such as Korea, Japan and China. C. lanceolata roots contain various chemical compounds including saponins like Panax ginseng. Although C. lanceolata are cultivated in different regions of South Korea, no variety has been developed. Therefore, it is necessary to develop discriminating methods such as molecular markers in C. lanceolata species. Methods and Results : To find simple sequence repeat (SSR) markers, we sequenced C. lanceolata genomic DNA using Illumina HiSeq 2000 System. A total of 250,455 putative SSR loci were obtained, and 26,334 non-redundant primers were designed to amplify these SSRs. Di-nucleotied repeats were the most abundant SSR reapeats, accounting for 89.53% (23,578) of primer designed SSRs. Tri-nucleotide, tetra-nucleotide and penta-nucleotide accounted for 8.44% (2,223), 1.3%, (348) 0.2% (55), respectively. Tri-, tetra-, and penta-nucleotide (total of 2,626 SSRs) were investigated in silico to identify polymorphism between individuals. Consequently, 573 SSRs showed polymorphism. Forty genomic SSR markers were tested in 16 C. lanceolata plants for determination of PCR amplification and polymorphism. From these primers, 27 (67.5%) amplified products and the average polymorphism information content (PIC) value was 0.52. Conclusion : We development 27 SSR markers from C. lanceolata using NGS, and it could be used for breeding of new varieties in the future.

Molecular targeting for the altered signaling pathways has been proven to be effective for the treatment of many types of human cancer, including colorectal cancer (CRC). The dual phosphatidylinositol-3-kinase (PI3K) and mammalian target of rapamycin (mTOR) inhibitor BEZ235 has shown to exhibit potent antitumor activity against solid tumors. Autophagy is a cellular lysosomal catabolic process to maintain metabolic homeostasis, which has been known to be induced in response to many therapeutic agents in cancer cells. This process is negatively regulated by mTOR and often acts as prosurvival or prodeath mechanism following cancer therapeutics. The current study was designed to investigate the antiproliferation activity of BEZ235 and to evaluate the role of autophagy induced by BEZ235 using HCT15 CRC cells bearing ras oncogene mutation. We found that BEZ235 decreases cell viability, which was mostly dependent on G1 arrest of cell cycle via suppression of cyclin A expression. BEZ235 affects PI3K/Akt/mTOR signaling pathway by increasing the phosphorylation of AKT at Ser473 and RAS/RAF/MEK/ERK pathway by decreasing the phosphorylation of ERK at Tyr204. BEZ235 also stimulated autophagy induction as evidenced by the increased expression of LC3-II and abundant acidic vesicular organelles (AVOs) in the cytoplasm. In addition, the combination of BEZ235 with autophagy inhibitor chloroquine, a known antagonist of autophagy, counteracted the antiproliferation effect of BEZ235. Thus, our study indicates that autophagy induced in response to BEZ235 treatment appears to act as cell death mechanism in HCT15 CRC cells.

This study was performed in order to provide basic data for predicting the usefulness of Jicama (Pachyrhizus erosus) as a food raw material. The changes in the physicochemical properties of freeze-dried and hot air-dried Jicama were investigated and analyzed. The moisture content of raw Jicama was 81.84%. The crude protein, crude fat, crude ash and carbohydrate content of hot air-dried Jicama powder were 2.85, 0.79, 7.93 and 88.44%, while those of freeze-dried Jicama powder were 3.93, 0.83, 7.92 and 87.32%, respectively on dry basis. Regarding the color values, the lightness of freeze-dried Jicama (92.86) was higher than that of the hot air-dried Jicama (88.01), whereas the redness (-0.67) and yellowness (3.21) of freeze-dried Jicama were lower than those of the hot air-dried Jicama (0.43) and (11.96), respectively. The brown index was lower in the freeze-dried Jicama (0.029) than in hot air-dried Jicama (0.107). The total sugar content showed no significant differences between freeze (46.49 mg/g) and hot air-dried Jicama (45.11 mg/g). Finally, the amylose content was higher in freeze-dried Jicama (5.66%) than in hot air-dried Jicama (6.63%).

This study assessed the fertilizing value of struvite deposit recovered from swine wastewater in cultivating Chinese cabbage. Struvite deposit was compared with commercial fertilizers: complex, organic and compost to evaluate the fertilizing effect of struvite deposit. Laboratory pot test obviously presented that the struvite deposit more facilitated the growth of Chinese cabbage than organic and compost fertilizers even though complex fertilizer was the most effective in growing Chinese cabbage. It was revealed that the growth rate of Chinese cabbage was simultaneously controlled by phosphorus (P) and potassium (K). Also, the nutrients such as nitrogen (N), P, K, calcium (Ca) and magnesium (Mg) were abundantly observed in the vegetable tissue of struvite pot. Specifically, P was the most abundant component in the vegetable tissue of struvite pot. Meanwhile, the utilization of struvite as a fertilizer led to the lower accumulation of chromium (Cr6+) than other pots, except for compost fertilizer pots, and no detection of cadmium (Cd), arsenic (As) and nickel (Ni) in the Chinese cabbage. The experimental results proved that the optimum struvite dosage for the cultivation of Chinese cabbage was 2.0 g struvite/kg soil. On the basis of these findings, it was concluded that the struvite deposits recovered from swine wastewater were effective as a multi-nutrient fertilizer for Chinese cabbage cultivation.

In this study, genetic diversity of wild Codonopsis lanceolata collected in Korea were analysed using SSR makers. Wild C. lanceolata roots were collected in Jeollanam-do Jangheung-gun Choentae Mountain as in roots. The wild C. lanceolata plants were cultivated in Chungbuk National University greenhouse and the leaves were sampled from 36 plants. The genomic DNA of C. lanceolata was extracted using CTAB. PCR was performed using a program of 35 cycles at 94℃ for 30 sec, 60℃ for 30 sec, and 72℃ for 30 sec with an pre-denaturation of 94℃ for 5 min and a final extension of 72℃ for 30 min. The PCR reaction mixture contains 5 pmole of primers and 20 ng of DNA template in a 20 μL reaction volume. The genotype of the analyzed samples were very different. Therefore, the wild C. lanceolata collected in Korea look genetically diverse.

Codonopsis lanceolata is a perennial climber. The roots are used as medicinal materials or vegetables. Recently, demand for C. lanceolata is increasing as a healthy food. C. lanceolata is distributed in India and East Asia such as China, Japan as well as Korea. In South Korea, this plant is widely cultivated in Gangwon-do province. No C. lanceolata varieties were developed in Korea. The objective of this study is to analyze genetic diversity of C. lanceolata cultivated in Korea using SSR makers. C. lanceolata roots were collected in each region were cultivated in Chungbuk National University greenhouse. Samples were obtained from fresh leaves of 5 plants from each collection region. The genomic DNA was extracted using CTAB. Genetic diversity was analysed using 4 sets of C. lanceolata SSR makers. PCR was performed in total 20 μL reaction volume containing 20 ng of DNA template, 5 pmole of primers. The genotypes of the analyzed samples were very similar. That means that the genetic diversity of C. lanceolata cultivated in Korea is very low.

We have generated 383 independent transgenic lines for genetically modified (GM) rice that contained PsGPD (Glyceraldehyde-3-Phosphate Dehydrogenase), ArCspA (Cold Shock Protein), BrTSR15 (Triple Stress Resistance 15) and BrTSR53 (Triple Stress Resistance 53) genes over-expression constructs under the control of the constitutive (CaMV 35S) promoter. TaqMan copy number assay was determined inserted T-DNA copy number. Also flanking sequence tags (FSTs) analysis was isolated from 203 single copy T-DNA lines of transgenic plants and sequence mapped to the rice chromosomes. In analyzing single copy lines, we identified 157 flanking sequence tags (FSTs), among which 58 (36%) were integrated into genic regions and 97 (62%) into intergenic regions. About 27 putative homozygous lines were obtained through multi-generations of planting, resistance screening and TaqMan copy number assay. To investigate the transgene expression patterns, quantitative real-time PCR analysis was performed using total RNAs from leaf tissue of single copy, intergenic region of T-DNA insertion and homozygous T2 plants. The mRNA expression levels of the examined transgenic rice were significantly increased in all of the transgenic plants. In addition, myc-tagged 35S:BrTSR15 and 35S:BrTSR53 transgenic plants were displayed higher levels of transgene protein. These results may be useful for producing of large-scale transgenic plants or T-DNA inserted mutants in rice.

Crops are exposed to various environmental stresses. These have been affecting the growth of crops, resulting in the severe loss of agronomic production in many countries. Therefore, development of new varieties of resistant crops is required to assure the desired productivity of crops in stress conditions. In this study, a putatively stress-related gene BrTSR53 was isolated from Brassica rapa. The BrTSR53 is 481 bp long and contains ORF region of 234 bp. The expression of BrTSR53 was determined by quantitative real-time PCR analysis. After 3 hr, the highest quantities of mRNA were revealed in cold and salt stress treatments. In drought stress treatments, there was the highest expression after 36 hr. Therefore, it was confirmed that the ORF in BrTSR53 should be a gene that confer increased resistance to B. rapa growing in different stress conditions. The ORF region of BrTSR53 gene was cloned into an expression vector, pYES-DEST52, and a new protein with molecular weight of 13 kDa was detected by western blot analysis. Also, stress tolerance tests showed that BrTSR53-ORF transgenic yeast exhibited increased resistance to the salt stresses compared with the control. In conclusion, the present data predicts that novel ORF in BrTSR53 can serve as an important genetic resource for abiotic stress resistance.

The purpose of development new variety ‘Miho’ (Hordeum vulgare L.) is a favorite with livestock feed and develop varieties resistant to disease and lodging. ‘Miho’ was carrying the growth habit of group Ⅲ, green and mid-wide leaf. Awn that are related to preference of livestock is a semi-smooth awn. This cultivar had 96cm of culm length, 650 of spikes per m2. Heading date of ‘Miho’ is April 27, and maturing dates on May 30, which were later than cultivar ‘Youngyang’. It also showed strong winter hardiness, and similar resistance to shattering and BaYMV compared with those of check one. The best thing among the traits of one is a new good quality with the plant green at the latter growing period. The average forage dry matter yield in the regional yield trial was about 13.1, 12.1 MT per ha in upland and paddy field, respectively, which were 9%, 2% higher than that of the check cultivar. It’s also showed 6.8% crude protein, 27.1% ADF (acid detergent fiber), and 67.5% TDN (Total Digestible Nutrients), including higher silage quality for whole crop barley. This cultivar would be suitable for the area whose daily minimum temperature was above -8℃ in January in Korean peninsula.

In this study, the experiment was carried out to produce methane by applying Semi-Continuous Leachate Recirculation Anaerobic Digestion System fed with source separated food waste from school cafeteria. There were two systems and each system consisted of a bioreactor and a leachate tank. Each bioreactor had a screen near the bottom of the reactor. 2L of Separated leachate was collected to the leachate tank each day by using a tubing pump and the leachate from the leachate tank was pumped to the bioreactor at the upper of the bioreactor. Through this circulation, the leachate having high concentration of VFAs was supplied to the bioreactor. At the beginning of the experiment, food waste/inoculum anaerobic sludge volume ratio was 2:8 that is 9g VS/L of OLR(Organic Loading Rate). Feeding was conducted every two weeks. Initial conditions of bioreactor was 30g VS/2･week and 33g VS/2･week were fed to bioreactor A and bioreactor B, respectively. Average biogas yields of the bioreactor were 0.723m³ Biogas/kg VS added in reactor A and 0.648m³ Biogas/kg VS added in reactor B.

Guppy has become a model organism for studying behavioral traits such as courtship and mate choice, as well as for understanding ecogeographic adaptation. Unfortunately, studying the early development of live bearers is more complicated than that of oviparous species, due to the inaccessibility of developing embryos for experimental manipulation. Ulrike et al. showed that the embryos could not be cultured for the entire period of their embryonic development. To optimize conditions embryo in vitro culture we established system for varying the concentration of fetal bovine serum in the medium impact on the embryonic development of in vitro embryos. For in vitro culture, embryos were incubated in 8 ml of sterile embryo medium (L-15 [Leibovitz] medium, supplemented with 5, 10, 15, and 20% fetal bovine serum respectively, 20 units/ml penicillin, and 200 mg/ml streptomycin) in a dark incubator at 25℃. Our study found that in 5% of FBS of the medium, embryos can be maintained until the middle-eyed. In 10% and 15% embryos can be maintained constant development; some of them can be fed; however, in 15% it is faster than 10%. And although in 20% of FBS can sustain rapid development of early stage, but ultimately died. According to our experimental data, both 10% and 15% FBS in medium can be used for in vitro culture, the slowly development in 10% FBS appears to be more conducive to observation.

Reactive oxygen species (ROS) are produced in organisms as the natural products of oxidative metabolism by environmental stress and pathogen invasion. ROS, such as superoxide anion and hydrogen peroxide, can be toxic to cells and tissues to cause oxidative stress. Recent study revealed that olive flounder (Paralichthys olivaceus) superoxide dismutase (SOD) has been identified as a partial gene and strongly induced to benzoin[a]pyrene and it was deduced indicator of aquatic oxidative stress responses, but its transcriptional response against viral infection has not been investigated. In the present study, spatial and temporal expression profile was analyzed to investigate the function of Of-SOD in the anti-viral response. Of-SOD transcripts were ubiquitously detected in diverse tissues with variable levels using a real-time PCR. The expression of Of-SOD was significantly higher in the muscle, liver and brain, but extremely low in the stomach and spleen. Following VHSV challenge, the expression of Of-SOD increased within 3 hours and subsequently decreased to the original level at 2 days post-challenge in kidney. Although expression pattern and induction time are slight differences depending on the tissue, the transcript of Of-SOD was consistently increased in acute infection response, but expression is low in the chronic response. Collectively, Of-SOD expressions were inducible after VHSV infection and they were probably involved in the immune response against viral challenge. These results suggest that SODs may play important roles in the immune defense system of P. olivaceus and perhaps contribute to the protective effects against oxidative stress in this flounder.

Molecular markers are useful for selecting to include superior character genetic like as strong immune system and rapid growth in fish. The marker is also very important part of breeding technology in Olive flounder (Paralichthys olivaceus). Single nucleotide polymorphisms (SNPs) marker is already in use widely for genomic research and breeding. But this SNPs marker hardly has been validated for screening functional genes in Olive flounder. We study identify single nucleotide polymorphisms (SNPs) on Expressed sequence tag (EST) database, develop usable SNP marker and apply to wild sample and cultured of olive flounder. As a result, Out of total 4.327 ESTs, 693contigs and 514 SNP from total contigs were detected while these substitutions include 297 transitions and 217 transversions. 144 developed markers were applied in 16 samples (wild 8, culture 8), Out of total marker, only 32 markers had detected polymorphic in sample. Polymorphism of 32 markers was observed in the variety genes region involved in immunity and protein synthesis. And the 32 marker were identified 21 transitions, 11 transversions, and indel was not detected in polymorphic SNPs. The analysis on heterozygosity by sample showed 0.34 in wild sample and 0.29 in cultured sample. In conclusion, we was identified SNP and Polymorphism by designed new marker, it supports that development marker is suitable for SNP detection and diversity analysis in Olive flounder. The outcome of this study can be basic data for researches for immunity gene and characteristic with SNP.

The genus Vicia comprises 166 annual or perennial species distributed mainly in Europe, Asia, and North America, also extending to the temperate regions of South America and tropical. However, utilization of SSR markers have not been investigated extensively in Vicia species as compared to other crop species. Here, we have assessed the potential for transferability (cross-species amplification) of cDNA microsatellites markers developed from common vetch (Vicia sativa subsp. sativa). In total, 226 alleles were detected in 36 microsatellite loci. The number of alleles per marker ranged from one to 20, with an average of 6.3. The gene diversity and polymorphism information content value averaged 0.540 and 0.503, with a range of 0–0.85 and 0–0.84 respectively. For transferability of the SSRs, amplification was carried out with selected from two to 8 accessions of 22 different Vicia species. For individual species, the successful amplification rate ranged from 32.6% in V. ervilia to 81.9% in V. sativa subsp. nigra, with average of 48.8%. As the rate of successful amplification of microsatellite markers generally correlates with genetic distance, these SSR markers are potentially useful in the analysis of genetic relationships between or within Vicia species.

We have generated 383 independent transgenic lines for genetically modified (GM) rice that contained GPD, UtrCSP, BrTSR15 and BrTSR53 genes overexpression constructs under the control of the constitutive CaMV 35S promoter. TaqMan copy number assay was determined inserted T-DNA copy number. Also FSTs analysis was isolated from 203 single copy T-DNA lines of transgenic plants and sequence mapped to the rice chromosomes. In analyzing single copy lines, we identified 95 FSTs, among which 37 (38.9%) were integrated into genic regions and 58 (61.1%) into intergenic regions. About 27 homozygous lines were obtained through multi-generations of planting, resistance screening and TaqMan copy number assay. To investigate the transgene expression patterns, quantitative real-time PCR analysis was performed using total RNAs from leaf tissue of single copy, intergenic region of T-DNA insertion and homozygous T2 plants. The mRNA expression levels of the examined transgenic rice were significantly increased in all of the transgenic plants. In addition, myc-tagged 35S::BrTSR15 and 35S::BrTSR53 transgenic plants were displayed higher levels of transgene protein than WT plants. These results may be useful for producing of large-scale transgenic plants or T-DNA inserted mutants in rice.

Fusarium head blight (FHB) is a major disease problem on wheat and barley around the world. F. graminearum produces trichothecenes mycotoxins such as nivalenol (NIV), deoxynivalenol (DON), and zearalenone (ZEA). The objectives of this study were to survey the natural occurrence of FHB and mycotoxins of 32 Korean wheat cultivars grown in 2011-2012 seasons at the National institute of crop science, Iksan, Korea. There was great deal of rainfall and high humidity during flowering time in May 2011. FHB incidence was counted by Fusarium infected spikes per square meter. The samples of 32 wheat cultivar were collected. The grain and flour samples were to analysis for DON and NIV by gas chromatography and ZEA by high performance liquid chromatography. The result showed that the average of FHB incidence(%) per square meter in 2011 and 2012 were 4.2%, 0.5% respectively. There were significant cultivar differences for FHB incidence ranged from 0% to 24% in 2011. All of 32 wheat cultivars contained 9-2088 ng/g for NIV and ten wheat cultivars contained 5.7-8.5 ng/g for ZEA. In addition, DON concentration of Tapdong, Shinmichal1, and Hanbaek were 217, 35 and 683 ng/g respectively. However, the grain and flour sample harvested in 2012 showed that lower FHB incidence and NIV concentration. These results showed that the 32 wheat cultivars harvested in 2011 were heavily contaminated with Fusarium mycotoxins (NIV, DON, ZEA).

With the purpose of improving ginsenoside production in Korean wild ginseng (Panax ginseng Meyer) mutant adventitious root lines, a synthetic gene encoding squalene synthase (PgSS2) was placed under the control of 35S promoter and transferred to Panax ginseng. Embryogenic callus obtained from ginseng adventitious root lines were transformed by infection with A. tumefaciens strain EHA105 containing the PgSS2 gene. Ten phosphinothricin-resistant plants were generated on selection medium, and the transgene integration and expression in these plants were confirmed by PAT test strip, RT-PCR and Southern hybridization. Ginsenoside analysis by HPLC revealed that the total contents of the 8 ginsenoside types (Rg1, Re, Rf, Rh1, Rb1, Rc, Rb2, Rd) in transgenic adventitious root lines were about 1.6-fold higher than that of the mutant control line (MCL1). This transformation method may facilitate the improvement of Panax ginseng in terms of the accumulation levels of ginsenoside.

This study was conducted to isolate a salt tolerant gene and to develop salt tolerant rice for reclaimed-saline areas through genetic transformation. A rice c/DRE binding factor 4(OsCBF4) cDNA was isolated from rice using RT-PCR. The full-length cDNA of the CBF4 gene consists of 1,429 nucleotides and 274 amino acid residues. In order to develop salt tolerant rice, transgenic rice plants containing the OsCBF4 gene were obtained via Agrobacterium-mediated transformation. The stable incorporation of the OsCBF4 gene into rice genome was confirmed by PCR and Southern analysis. The stable expression of introduced gene was also validated by RT-PCR analysis in T2 plants. Biological assay of T3 progeny of the transgenic plants in Yoshida solution containing 120mM Nacl for 2weeks, confirmed that the OsCBF4 confers salt tolerance to transgenic rice plants. OsCBF4 transgene in the transgenic line CBF4-10 was markedly expressed up to over three-fold in the leaf by 120 mM NaCl treatment. Real-time PCR analysis revealed that the levels of the transgene expression were markedly increased under salt treatment. The transgenic line CBF4-10 which showed highest ability to recover from the saline stress could be used as a potential source for salt tolerance in rice breeding programs

Sequence diversity was accumulated through evolution and breeding process. A set of 595 PCR-based novel insertion/deletion (InDel) markers was designed in order to widen the genetic basis for national rice breeding programs. The markers were generated by analyzing of 40 Korean cultivars and published genome sequences of rice(Oryza sativa L. spp japonica). We selected 112 markers spread across all rice chromosomes among the 595 InDel markers, and they showed polymorphic between rice cultivars, which are 284 Korean japonica and Tongil varieties. Due to their simplicity in design and robustness in genotyping, these InDel markers have been routinely used in quantitative trait loci (QTL) mapping studies and marker assisted selection programs for rice. Moreover, the PCR amplification type of InDel markers was converged to digital code, 0 or 1 and then finally represented as one- and two dimensional bar-code system, which could easily differentiate genetically highly homologous japonica rice cultivars. The developed InDel markers uniquely discriminated among each of the Korean cultivars. Therefore, the systems we developed may be valuable tools in discrimination from cultivars

Mungbean (Vigna radiata) is a fast-growing, warm-season legume crop that is primarily cultivated in developing countries of Asia. We constructed a draft genome sequence of mungbean to facilitate genome research into the subgenus Ceratotropis and to enable a better understanding of the evolution of leguminous species. The draft genome sequence covers 80% of the estimated genome, of which 50.1% consists of repetitive sequences. In total, 22,427 high confidence protein-coding genes were predicted. Based on the de novo assembly of additional wild mungbean species, the divergence of what was eventually domesticated and the sampled wild mungbean species appears to have predated domestication. Moreover, the de novo assembly of a tetraploid Vigna species (Vigna reflexo-pilosa var. glabra) provided genomic evidence of a recent allopolyploid event. To further study speciation, we compared de novo RNA-seq assemblies of 22 accessions of 18 Vigna species and protein sets of Glycine max and Cajanus cajan. The species tree was constructed by a Bayesian Markov chain Monte Carlo method using highly confident orthologs shared by all 24 accessions. The present assembly of V. radiata var. radiata will facilitate genome research and accelerate molecular breeding of the subgenus Ceratotropis.