Sphingomonas cannabina sp. nov., isolated from Cannabis sativa L. 'Cheungsam'.

A Gram-negative, aerobic, rod-shaped bacterium, designated DM2-R-LB4T was isolated from Cannabis sativa L. 'Cheungsam' in Andong, Republic of Korea. The strain DM2-R-LB4T grew at temperatures of 15-45 °C (optimum, 30-37 °C), pH of 5.5-9 (optimum, 8.0), and 0-2 % (w/v) NaCl concentration (optimum, 0%). Phylogenetic analyses based on the 16S rRNA gene sequences revealed that strain DM2-R-LB4T is related to species of the genus Sphingomonas, and shared 97.8 and 97.5% similarity to Sphingomonas kyenggiensis KCTC 42244T and Sphingomonas leidyi DSM 4733T, respectively. The DNA G+C content was 67.9 mol% and genome analysis of the strain DM2-R-LB4T revealed that the genome size was 4 386 171 bp and contained 4 009 predicted protein-coding genes. The average nucleotide identity (ANI) values between strain DM2-R-LB4T and S. kyenggiensis KCTC 42244T, and S. leidyi DSM 4733T was 76.8 and 76.7 %, respectively, while the values of digital DNA-DNA hybridization (dDDH) were 20.7 and 20.6 %, respectively. C14 : 0 2-OH, C16 : 0, and summed feature 8 (C18 : 1 ω6c and/or C18 : 1 ω7c) were the major fatty acids (>10 %) in the strain DM2-R-LB4T. The polar lipids comprised diphosphatidylglycerol (DPG), phosphatidylethanolamine (PE), phosphatidylglycerol (PG), phosphatidylcholine (PC), sphingoglycolipid (SGL), glycolipid (GL), phospholipid (PL), and two unidentified polar lipids (L1 and L2). Ubiquinone-10 (Q-10) was the only respiratory quinone. The polyamine pattern was found to contain homospermidine, putrescine, and spermidine. The results of phylogenetic anlayses, polyphasic studies, revealed that strain DM2-R-LB4T represents a novel species of the genus Sphingomonas, for which the name Sphingomonas cannabina sp. nov., is proposed. The type strain is DM2-R-LB4T (=KCTC 92075T = GDMCC 1.3018T).

Highly Efficient Bioconversion of trans-Resveratrol to δ-Viniferin Using Conditioned Medium of Grapevine Callus Suspension Cultures.

δ-Viniferin is a resveratrol dimer that possesses potent antioxidant properties and has attracted attention as an ingredient for cosmetic and nutraceutical products. Enzymatic bioconversion and plant callus and cell suspension cultures can be used to produce stilbenes such as resveratrol and viniferin. Here, δ-viniferin was produced by bioconversion from trans-resveratrol using conditioned medium (CM) of grapevine (Vitis labruscana) callus suspension cultures. The CM converted trans-resveratrol to δ-viniferin immediately after addition of hydrogen peroxide (H2O2). Peroxidase activity and bioconversion efficiency in CM increased with increasing culture time. Optimized δ-viniferin production conditions were determined regarding H2O2 concentration, incubation time, temperature, and pH. Maximum bioconversion efficiency reached 64% under the optimized conditions (pH 6.0, 60 °C, 30 min incubation time, 6.8 mM H2O2). In addition, in vitro bioconversion of trans-resveratrol was investigated using CM of different callus suspension cultures, showing that addition of trans-resveratrol and H2O2 to the CM led to production of δ-viniferin via extracellular peroxidase-mediated oxidative coupling of two molecules of trans-resveratrol. We thus propose a simple and low-cost method of δ-viniferin production from trans-resveratrol using CM of plant callus suspension cultures, which may constitute an alternative approach for in vitro bioconversion of valuable molecules.

Mutation of GmIPK1 Gene Using CRISPR/Cas9 Reduced Phytic Acid Content in Soybean Seeds.

Phytic acid (PA) acts as an antinutrient substance in cereal grains, disturbing the bioavailability of micronutrients, such as iron and zinc, in humans, causing malnutrition. GmIPK1 encodes the inositol 1,3,4,5,6-pentakisphosphate 2-kinase enzyme, which converts myo-inopsitol-1,3,4,5,6-pentakisphosphate (IP5) to myo-inositol-1,2,3,4,5,6-hexakisphosphate (IP6) in soybean (Glycine max L.). In this study, for developing soybean with low PA levels, we attempted to edit the GmIPK1 gene using the CRISPR/Cas9 system to introduce mutations into the GmIPK1 gene with guide RNAs in soybean (cv. Kwangankong). The GmIPK1 gene was disrupted using the CRISPR/Cas9 system, with sgRNA-1 and sgRNA-4 targeting the second and third exon, respectively. Several soybean Gmipk1 gene-edited lines were obtained in the T0 generation at editing frequencies of 0.1-84.3%. Sequencing analysis revealed various indel patterns with the deletion of 1-9 nucleotides and insertions of 1 nucleotide in several soybean lines (T0). Finally, we confirmed two sgRNA-4 Gmipk1 gene-edited homozygote soybean T1 plants (line #21-2: 5 bp deletion; line #21-3: 1 bp insertion) by PPT leaf coating assay and PCR analysis. Analysis of soybean Gmipk1 gene-edited lines indicated a reduction in PA content in soybean T2 seeds but did not show any defects in plant growth and seed development.

Pedobacter endophyticus sp. nov., an endophytic bacterium isolated from Carex pumila.

An aerobic, Gram-stain-negative, weak-motile, short-rod-shaped bacterial strain, designated JBR3-12T, was isolated from halophyte Carex pumila plants, and its taxonomic position was investigated by using a polyphasic taxonomic approach. The strain produced a pink pigment on tryptic soy agar and grew optimally at 25 °C, pH 8 and in the presence of 3 % (w/v) NaCl. Results of phylogenetic analysis based on 16S rRNA gene sequences showed that strain JBR3-12T formed a lineage within the genus Pedobacter and was most closely related to Pedobacter sandarakinus DS-27T (98.0 %) and Pedobacter agri PB92T (97.6 %). The DNA G+C content of the genome was 41.3 mol%; the whole genome length was 5 426 070 bp. The major fatty acids of JBR3-12T were iso-C15 : 0, summed feature 3 (comprising C16 : 1 ω6c and/or C16 : 1 ω7c) and iso-C17 : 0 3-OH. The predominant polar lipid was phosphatidylethanolamine. The predominant quinone was menaquinone-7. Based on its phenotypic, phylogenetic and genotypic features, strain JBR3-12T is proposed to represent a novel species of the genus Pedobacter, for which the name is Pedobacter endophyticus sp. nov. The type strain is JBR3-12T (=KCTC 82363T=NBRC 114901T).

Synergistic Effect of Methyl Jasmonate and Abscisic Acid Co-Treatment on Avenanthramide Production in Germinating Oats.

The oat (Avena sativa L.) is a grain of the Poaceae grass family and contains many powerful anti-oxidants, including avenanthramides as phenolic alkaloids with anti-inflammatory, anti-oxidant, anti-itch, anti-irritant, and anti-atherogenic activities. Here, the treatment of germinating oats with methyl jasmonate (MeJA) or abscisic acid (ABA) resulted in 2.5-fold (582.9 mg/kg FW) and 2.8-fold (642.9 mg/kg FW) increase in avenanthramide content, respectively, relative to untreated controls (232.6 mg/kg FW). Moreover, MeJA and ABA co-treatment synergistically increased avenanthramide production in germinating oats to 1505 mg/kg FW. Individual or combined MeJA and ABA treatment increased the expression of genes encoding key catalytic enzymes in the avenanthramide-biosynthesis pathway, including hydroxycinnamoyl-CoA:hydrocyanthranilate N-hydroxycinnamoyl transferase (HHT). Further analyses showed that six AsHHT genes were effectively upregulated by MeJA or ABA treatment, especially AsHHT4 for MeJA and AsHHT5 for ABA, thereby enhancing the production of all three avenanthramides in germinating oats. Specifically, AsHHT5 exhibited the highest expression following MeJA and ABA co-treatment, indicating that AsHHT5 played a more crucial role in avenanthramide biosynthesis in response to MeJA and ABA co-treatment of germinating oats. These findings suggest that elicitor-mediated metabolite farming using MeJA and ABA could be a valuable method for avenanthramide production in germinating oats.

The Effect of Sodium Butyrate on Adventitious Shoot Formation Varies among the Plant Species and the Explant Types.

Histone acetylation plays an important role in plant growth and development. Here, we investigated the effect of sodium butyrate (NaB), a histone deacetylase inhibitor, on adventitious shoot formation from protoplast-derived calli and cotyledon explants of tobacco (Nicotiana benthamiana) and tomato (Solanum lycopersicum). The frequency of adventitious shoot formation from protoplast-derived calli was higher in shoot induction medium (SIM) containing NaB than in the control. However, the frequency of adventitious shoot formation from cotyledon explants of tobacco under the 0.1 mM NaB treatment was similar to that in the control, but it decreased with increasing NaB concentration. Unlike in tobacco, NaB decreased adventitious shoot formation in tomato explants in a concentration-dependent manner, but it did not have any effect on adventitious shoot formation in calli. NaB inhibited or delayed the expression of D-type cyclin (CYCD3-1) and shoot-regeneration regulatory gene WUSCHEL (WUS) in cotyledon explants of tobacco and tomato. However, compared to that in control SIM, the expression of WUS was promoted more rapidly in tobacco calli cultured in NaB-containing SIM, but the expression of CYCD3-1 was inhibited. In conclusion, the effect of NaB on adventitious shoot formation and expression of CYCD3-1 and WUS genes depended on the plant species and whether the effects were tested on explants or protoplast-derived calli.

The Synergistic Effect of Co-Treatment of Methyl Jasmonate and Cyclodextrins on Pterocarpan Production in Sophora flavescens Cell Cultures.

Pterocarpans are derivatives of isoflavonoids, found in many species of the family Fabaceae. Sophora flavescens Aiton is a promising traditional Asian medicinal plant. Plant cell suspension cultures represent an excellent source for the production of valuable secondary metabolites. Herein, we found that methyl jasmonate (MJ) elicited the activation of pterocarpan biosynthetic genes in cell suspension cultures of S. flavescens and enhanced the accumulation of pterocarpans, producing mainly trifolirhizin, trifolirhizin malonate, and maackiain. MJ application stimulated the expression of structural genes (PAL, C4H, 4CL, CHS, CHR, CHI, IFS, I3'H, and IFR) of the pterocarpan biosynthetic pathway. In addition, the co-treatment of MJ and methyl-ß-cyclodextrin (MeßCD) as a solubilizer exhibited a synergistic effect on the activation of the pterocarpan biosynthetic genes. The maximum level of total pterocarpan production (37.2 mg/g dry weight (DW)) was obtained on day 17 after the application of 50 µM MJ on cells. We also found that the combined treatment of cells for seven days with MJ and MeßCD synergistically induced the pterocarpan production (trifolirhizin, trifolirhizin malonate, and maackiain) in the cells (58 mg/g DW) and culture medium (222.7 mg/L). Noteworthy, the co-treatment only stimulated the elevated extracellular production of maackiain in the culture medium, indicating its extracellular secretion; however, its glycosides (trifolirhizin and trifolirhizin malonate) were not detected in any significant amounts in the culture medium. This work provides new strategies for the pterocarpan production in plant cell suspension cultures, and shows MeßCD to be an effective solubilizer for the extracellular production of maackiain in the cell cultures of S. flavescens.

Neobacillus endophyticus sp. nov., an endophytic bacterium isolated from Selaginella involvens roots.

A Gram-stain-positive, facultatively anaerobic, rod-shaped, endospore-forming, oxidase-positive, and catalase-negative strain designated as BRMEA1T was isolated from the surface-sterilized Selaginella involvens roots. Growth of strain BRMEA1T was found to occur at pH 6.0-8.0 (optimum, pH 7.0), 15-50 °C (optimum, 25-30 °C) and in the absence of NaCl. Phylogenetic analysis based on 16S rRNA gene sequences indicated that strain BRMEA1T formed a lineage within the genus Neobacillus (family Bacillaceae) and showed the highest sequence similarity to Neobacillus drentensis DSM 15600T (98.3 %) and Neobacillus fumarioli KCTC 13885T (98.2 %), and less than 98.2 % 16S rRNA gene sequence similarity to the other members of the genus Neobacillus. Whole-genome analysis of strain BRMEA1T comprised a circular chromosome (5 632 809 bp in size) with 38.5 mol% G+C content. Digital DNA-DNA hybridization analyses revealed that strain BRMEA1T showed 20.5 and 22.0% genomic DNA relatedness with the closest species, N. drentensis DSM 15600T and N. fumarioli KCTC 13885T, respectively. The whole-genome sequence of strain BRMEA1T showed the presence of 11 specific conserved signature indels for the genus Neobacillus. The major cellular fatty acids (>10 %) of strain BRMEA1T were found to be iso-C15 : 0 and anteiso-C15 : 0, while the major polar lipids were found to be diphosphatidylglycerol, phosphatidylethanolamine and phosphatidylglycerol. Polyphasic analysis results revealed that BRMEA1T represents a novel species of the genus Neobacillus, with the proposed name Neobacillus endophyticus sp. nov. The type strain is BRMEA1T (=KCTC 43208T=CCTCC AB 2020071T).

Plant synthetic GP4 and GP5 proteins from porcine reproductive and respiratory syndrome virus elicit immune responses in pigs.

MAIN CONCLUSION: We demonstrated successful overexpression of porcine reproductive and respiratory syndrome virus (PRRSV)-derived GP4D and GP5D antigenic proteins in Arabidopsis. Pigs immunized with transgenic plants expressing GP4D and GP5D proteins generated both humoral and cellular immune responses to PRRSV. Porcine reproductive and respiratory syndrome virus (PRRSV) causes PRRS, the most economically significant disease affecting the swine industry worldwide. However, current commercial PRRSV vaccines (killed virus or modified live vaccines) show poor efficacy and safety due to concerns such as reversion of virus to wild type and lack of cross protection. To overcome these problems, plants are considered a promising alternative to conventional platforms and as a vehicle for large-scale production of recombinant proteins. Here, we demonstrate successful production of recombinant protein vaccine by expressing codon-optimized and transmembrane-deleted recombinant glycoproteins (GP4D and GP5D) from PRRSV in planta. We generated transgenic Arabidopsis plants expressing GP4D and GP5D proteins as candidate antigens. To examine immunogenicity, pigs were fed transgenic Arabidopsis leaves expressing the GP4D and GP5D antigens (three times at 2-week intervals) and then challenged with PRRSV at 6-week post-initial treatment. Immunized pigs showed significantly lower lung lesion scores and reduced viremia and viral loads in the lung than pigs fed Arabidopsis leaves expressing mYFP (control). Immunized pigs also had higher titers of PRRSV-specific antibodies and significantly higher levels of pro-inflammatory cytokines (TNF-α and IL-12). Furthermore, the numbers of IFN-γ+-producing cells were higher, and those of regulatory T cells were lower, in GP4D and GP5D immunized pigs than in control pigs. Thus, plant-derived GP4D and GP5D proteins provide an alternative platform for producing an effective subunit vaccine against PRRSV.

Down-regulation of GIGANTEA-like genes increases plant growth and salt stress tolerance in poplar.

The flowering time regulator GIGANTEA (GI) connects networks involved in developmental stage transitions and environmental stress responses in Arabidopsis. However, little is known about the role of GI in growth, development and responses to environmental challenges in the perennial plant poplar. Here, we identified and functionally characterized three GI-like genes (PagGIa, PagGIb and PagGIc) from poplar (Populus alba × Populus glandulosa). PagGIs are predominantly nuclear localized and their transcripts are rhythmically expressed, with a peak around zeitgeber time 12 under long-day conditions. Overexpressing PagGIs in wild-type (WT) Arabidopsis induced early flowering and salt sensitivity, while overexpressing PagGIs in the gi-2 mutant completely or partially rescued its delayed flowering and enhanced salt tolerance phenotypes. Furthermore, the PagGIs-PagSOS2 complexes inhibited PagSOS2-regulated phosphorylation of PagSOS1 in the absence of stress, whereas these inhibitions were eliminated due to the degradation of PagGIs under salt stress. Down-regulation of PagGIs by RNA interference led to vigorous growth, higher biomass and enhanced salt stress tolerance in transgenic poplar plants. Taken together, these results indicate that several functions of Arabidopsis GI are conserved in its poplar orthologues, and they lay the foundation for developing new approaches to producing salt-tolerant trees for sustainable development on marginal lands worldwide.

A vacuolar ß-glucosidase homolog that possesses glucose-conjugated abscisic acid hydrolyzing activity plays an important role in osmotic stress responses in Arabidopsis.

The phytohormone abscisic acid (ABA) plays a critical role in various physiological processes, including adaptation to abiotic stresses. In Arabidopsis thaliana, ABA levels are increased both through de novo biosynthesis and via ß-glucosidase homolog1 (BG1)-mediated hydrolysis of Glc-conjugated ABA (ABA-GE). However, it is not known how many different ß-glucosidase proteins produce ABA from ABA-GE and how the multiple ABA production pathways are coordinated to increase ABA levels. Here, we report that a previously undiscovered ß-glucosidase homolog, BG2, produced ABA by hydrolyzing ABA-GE and plays a role in osmotic stress response. BG2 localized to the vacuole as a high molecular weight complex and accumulated to high levels under dehydration stress. BG2 hydrolyzed ABA-GE to ABA in vitro. In addition, BG2 increased ABA levels in protoplasts upon application of exogenous ABA-GE. Overexpression of BG2 rescued the bg1 mutant phenotype, as observed for the overexpression of NCED3 in bg1 mutants. Multiple Arabidopsis bg2 alleles with a T-DNA insertion in BG2 were more sensitive to dehydration and NaCl stress, whereas BG2 overexpression resulted in enhanced resistance to dehydration and NaCl stress. Based on these observations, we propose that, in addition to the de novo biosynthesis, ABA is produced in multiple organelles by organelle-specific ß-glucosidases in response to abiotic stresses.

Overexpression of the IbMYB1 gene in an orange-fleshed sweet potato cultivar produces a dual-pigmented transgenic sweet potato with improved antioxidant activity.

The R2R3-type protein IbMYB1 is a key regulator of anthocyanin biosynthesis in the storage roots of sweet potato [Ipomoea batatas (L.) Lam]. Previously, we demonstrated that IbMYB1 expression stimulated anthocyanin pigmentation in tobacco leaves and Arabidopsis. Here, we generated dual-pigmented transgenic sweet potato plants that accumulated high levels of both anthocyanins and carotenoids in a single sweet potato storage root. An orange-fleshed cultivar with high carotenoid levels was transformed with the IbMYB1 gene under the control of either the storage root-specific sporamin 1 (SPO1) promoter or the oxidative stress-inducible peroxidase anionic 2 (SWPA2) promoter. The SPO1-MYB transgenic lines exhibited higher anthocyanin levels in storage roots than empty vector control (EV) or SWPA2-MYB plants, but carotenoid content was unchanged. SWPA2-MYB transgenic lines exhibited higher levels of both anthocyanin and carotenoids than EV plants. Analysis of hydrolyzed anthocyanin extracts indicated that cyanidin and peonidin predominated in both overexpression lines. Quantitative reverse transcription-polymerase chain reaction analysis demonstrated that IbMYB1 expression in both IbMYB1 transgenic lines strongly induced the upregulation of several genes in the anthocyanin biosynthetic pathway, whereas the expression of carotenoid biosynthetic pathway genes varied between transgenic lines. Increased anthocyanin levels in transgenic plants also promoted the elevation of proanthocyanidin and total phenolic levels in fresh storage roots. Consequently, all IbMYB1 transgenic plants displayed much higher antioxidant activities than EV plants. In field cultivations, storage root yields varied between the transgenic lines. Taken together, our results indicate that overexpression of IbMYB1 is a highly promising strategy for the generation of transgenic plants with enhanced antioxidant capacity.

Differential responses of three sweetpotato metallothionein genes to abiotic stress and heavy metals.

Metallothioneins (MTs) are cysteine-rich, low molecular weight, metal-binding proteins that are widely distributed in living organisms. Plants produce metal-chelating proteins such as MTs to overcome the toxic effects of heavy metals. We cloned three MT genes from sweetpotato leaves [Ipomoea batatas (L.) Lam.]. The three IbMT genes were classified according to their cysteine residue alignment into type 1 (IbMT1), type 2 (IbMT2), and type 3 (IbMT3). IbMT1 was the most abundantly transcribed MT. It was predominantly expressed in leaves, roots, and callus. IbMT2 transcript was detected only in stems and fibrous roots, whereas IbMT3 was strongly expressed in leaves and stems. The IbMT expression profiles were investigated in plants exposed to heavy metals and abiotic stresses. The levels of IbMT1 expression were strongly elevated in response to Cd and Fe, and moderately higher in response to Cu. The IbMT3 expression pattern in response to heavy metals was similar to that of IbMT1. Exposure to abiotic stresses such as methyl viologen (MV; paraquat), NaCl, polyethylene glycol (PEG), and H2O2 up-regulated IbMT expression; IbMT1 responded strongly to MV and NaCl, whereas IbMT3 was induced by low temperature and PEG. Transgenic Escherichia coli overexpressing IbMT1 protein exhibited results suggest that IbMT could be a useful tool for engineering plants with enhanced tolerance to environmental stresses and heavy metals.

Down-regulation of sweetpotato lycopene ß-cyclase gene enhances tolerance to abiotic stress in transgenic calli.

Lycopene ß-cyclase (LCY-ß) is a key enzyme involved in the synthesis of α- and ß-branch carotenoids such as α-carotene and ß-carotene through the cyclization of lycopene. IbLCY-ß had a length of 1,506 bp and approximately 80 % nucleotide sequence identity with that of tomato LCY-ß. IbLCY-ß was strongly expressed in leaves, and expression was enhanced by salt-stress and osmotic-stress conditions. To characterize the LCY-ß gene (IbLCY-ß) of sweetpotato (Ipomoea batatas), it was isolated and transformed into calli of white-fleshed sweetpotato using an IbLCY-ß-RNAi vector. Transgenic IbLCY-ß-RNAi calli had yellow to orange color and higher antioxidant activity compared to that of white, nontransgenic (NT) calli. Transgenic cells had significantly higher contents of total carotenoids, although lycopene was not detected in transgenic or NT cells. All transgenic calli had strongly activated expression of carotenoid biosynthetic genes such as ß-carotene hydroxylases (CHY-ß), cytochrome P450 monooxygenases (P450), and carotenoid cleavage dioxigenase 1 (CCD1). Transgenic cells exhibited less salt-induced oxidative-stress damage compared to that of NT cells, and also had greater tolerance for polyethylene glycol (PEG)-mediated drought compared to that of NT cells, due to the higher water content and reduced malondialdehyde (MDA) content. The abscisic acid content was also higher in transgenic cells. These results show that a study of IbLCY-ß can facilitate understanding of the carotenoid biosynthetic pathway in sweetpotato. IbLCY-ß could be useful for developing transgenic sweetpotato enriched with nutritional carotenoids and with greater tolerance to abiotic stresses.

Downregulation of the lycopene ε-cyclase gene increases carotenoid synthesis via the ß-branch-specific pathway and enhances salt-stress tolerance in sweetpotato transgenic calli.

Lycopene ε-cyclase (LCY-ε) is involved in the first step of the α-branch synthesis pathway of carotenoids from lycopene in plants. In this study, to enhance carotenoid synthesis via the ß-branch-specific pathway [which yields ß-carotene and abscisic acid (ABA)] in sweet potato, the expression of IbLCY-ε was downregulated by RNAi (RNA interference) technology. The RNAi-IbLCY-ε vector was constructed using a partial cDNA of sweet potato LCY-ε isolated from the storage root and introduced into cultured sweet potato cells by Agrobacterium-mediated transformation. Both semi-quantitative Reverse transcription polymerase chain reaction (RT-PCR) of carotenoid biosynthesis genes and high-performance liquid chromatography (HPLC) analysis of the metabolites in transgenic calli, in which the LCY- εgene was silenced, showed the activation of ß-branch carotenoids and its related genes. In the transgenic calli, the ß-carotene content was approximately 21-fold higher than in control calli, whereas the lutein content of the transgenic calli was reduced to levels undetectable by HPLC. Similarly, expression of the RNAi-IbLCY-ε transgene resulted in a twofold increase in ABA content compared to control calli. The transgenic calli showed significant tolerance of 200 mM NaCl. Furthermore, both the ß-branch carotenoids content and the expression levels of various branch-specific genes were higher under salt stress than in control calli. These results suggest that, in sweet potato, downregulation of the ε-cyclization of lycopene increases carotenoid synthesis via the ß-branch-specific pathway and may positively regulate cellular defenses against salt-mediated oxidative stress.

Expression of the sweetpotato R2R3-type IbMYB1a gene induces anthocyanin accumulation in Arabidopsis.

R2R3-type MYB transcription factors (TFs) play important roles in transcriptional regulation of anthocyanins. The R2R3-type IbMYB1 is known to be a key regulator of anthocyanin biosynthesis in the storage roots of sweetpotato. We previously showed that transient expression of IbMYB1a led to anthocyanin pigmentation in tobacco leaves. In this article, we generated transgenic Arabidopsis plants expressing the IbMYB1a gene under the control of CaMV 35S promoter, and the sweetpotato SPO and SWPA2 promoters. Overexpression of IbMYBa in transgenic Arabidopsis produced strong anthocyanin pigmentation in seedlings and generated a deep purple color in leaves, stems and seeds. Reverse transcription-polymerase chain reaction analysis showed that IbMYB1a expression induced upregulation of several structural genes in the anthocyanin biosynthetic pathway, including 4CL, CHI, F3'H, DFR, AGT, AAT and GST. Furthermore, overexpression of IbMYB1a led to enhanced expression of the AtTT8 (bHLH) and PAP1/AtMYB75 genes. high-performance liquid chromatography analysis revealed that IbMYB1a expression led to the production of cyanidin as a major core molecule of anthocyanidins in Arabidopsis, as occurs in the purple leaves of sweetpotato (cv. Sinzami). This result shows that the IbMYB1a TF is sufficient to induce anthocyanin accumulation in seedlings, leaves, stems and seeds of Arabidopsis plants.

Comparative characterization of sweetpotato antioxidant genes from expressed sequence tags of dehydration-treated fibrous roots under different abiotic stress conditions.

Drought stress is one of the most adverse conditions for plant growth and productivity. The plant antioxidant system is an important defense mechanism and includes antioxidant enzymes and low-molecular weight antioxidants. Understanding the biochemical and molecular responses to drought is essential for improving plant resistance to water-limited conditions. Previously, we isolated and characterized expressed sequence tags (ESTs) from a full-length enriched cDNA library prepared from fibrous roots of sweetpotato subjected to dehydration stress (Kim et al. in BMB Rep 42:271-276, [5]). In this study, we isolated and characterized 11 sweetpotato antioxidant genes from sweetpotato EST library under various abiotic stress conditions, which included six intracellular CuZn superoxide dismutases (CuZnSOD), ascorbate peroxidase, catalase, glutathione peroxidase (GPX), glutathione-S-transferase, thioredoxin (TRX), and five extracellular peroxidase genes. The expression of almost all the antioxidant genes induced under dehydration treatments occurred in leaves, with the exception of extracellular swPB6, whereas some antioxidant genes showed increased expression levels in the fibrous roots, such as intracellular GPX, TRX, extracellular swPA4, and swPB7 genes. During various abiotic stress treatments in leaves, such as exposure to NaCl, cold, and abscisic acid, several intracellular antioxidant genes were strongly expressed compared with the expression of extracellular antioxidant genes. These results indicated that some intracellular antioxidant genes, especially swAPX1 and CuZnSOD, might be specifically involved in important defense mechanisms against oxidative stress induced by various abiotic stresses including dehydration in sweetpotato plants.

Effects of Carbohydrates on Rosmarinic Acid Production and In Vitro Antimicrobial Activities in Hairy Root Cultures of Agastache rugosa.

Agastache rugosa (popularly known as Korean mint) belongs to the Lamiaceae family and comprises 22 species of perennial aromatic medicinal species native to East Asian countries, such as Korea, Taiwan, Japan, and China. A. rugosa contains many phenolic compounds that exhibit pharmacological and physiological activities, including antioxidant, anticancer, antiviral, antifungal, and antibacterial activities. The highest concentrations of rosmarinic acid and its isomers have been reported in the roots of A. rugosa. In this in vitro study, hairy roots of A. rugosa were obtained and the carbohydrates (sorbitol, mannitol, glucose, maltose, galactose, mannose, and sucrose) were evaluated to determine those that were optimal for rosmarinic acid production and hairy root growth. Antioxidant and antibacterial activities of extracts of A. rugosa were also assessed. The best carbon source for A. rugosa hairy root cultures was sucrose, considering biomass productivity (0.460 ± 0.034 mg/30 mL), rosmarinic acid production (7.656 ± 0.407 mg/g dry weight), and total phenolic content (12.714 ± 0.202 mg/g gallic acid equivalent). Antioxidant and antimicrobial activities were displayed by A. rugosa hairy roots cultured in liquid medium supplemented with 100 mM sucrose. Twenty-five bacterial strains, including multidrug-resistant bacteria and one pathogenic yeast strain, were used for antimicrobial screening of A. rugosa hairy roots. The hairy root extracts displayed antibacterial activity against Micrococcus luteus (KCTC 3063) and Bacillus cereus (KCTC 3624). The inhibition of these bacteria was greater using A. rugosa hairy roots with the highest levels of phenolic compounds cultured in the presence of sucrose, compared to hairy roots with the lowest levels of phenolic compounds cultured in the presence of fructose. Considering hairy root biomass, phenolic compound production, and antibacterial activity, sucrose is the best carbon source for A. rugosa hairy root cultures.

Bioplastic (poly-3-hydroxybutyrate)-producing Massilia endophytica sp. nov., isolated from Cannabis sativa L. 'Cheungsam'.

A rod-shaped, motile, Gram-negative bacterial strain named DM-R-R2A-13T was isolated from the plant Cannabis sativa L. 'Cheungsam'. The phylogenetic analysis of the 16S rRNA gene sequence revealed that strain DM-R-R2A-13T belongs to the family Oxalobacteraceae and is closely related to members of the genus Massilia, with Massilia flava (97.58% sequence similarity) and Massilia armeniaca (97.37% sequence similarity) being the closest members. The digital DNA-DNA hybridization (dDDH) values between strain DM-R-R2A-13T and Massilia flava CGMCC 1.10685T and Massilia armeniaca ZMN-3Twere 22.2% and 23.3%, while the average nucleotide identity (ANI) values were 78.85% and 79.63%, respectively. The DNA G+C content was measured to be 64.6 mol%. Moreover, the bacterium was found to contain polyhydroxyalkanoate (PHA) granules based on transmission electron microscopy, indicating its potential to produce bioplastic. Genome annotation revealed the presence of PHA synthase genes (phaC, phaR, phaP, and phaZ), and the biopolymer was identified as poly-3-hydroxybutyrate (PHB) based on nuclear magnetic resonance (NMR) and Fourier transform infrared spectroscopy (FTIR) analyses. Using maltose as a carbon source, the strain produced PHB of up to 58.06% of its dry cell weight. Based on the phenotypic, chemotaxonomic, and phylogenetic characteristics, it has been determined that DM-R-R2A-13T represents a novel species belonging to the genus Massilia. As such, the name Massilia endophytica sp. nov. is proposed for this newly identified species. The type strain is DM-R-R2A-13T (= KCTC 92072T = GDMCC 1.2920T).

Fabrication of molded chalcogenide-glass lens for thermal imaging applications.

With the recent development of less costly uncooled detector technology, expensive optics are among the remaining significant cost drivers. As a potential solution to this problem, the fabrication of IR lenses using chalcogenide glass has been studied in recent years. We report on the fabrication of a molded chalcogenide-glass lens for car night vision and on the evaluation of the lens. The moldability of chalcogenide glass was characterized through transcription properties of the mold's surface. In addition, both IR transmittance and x-ray diffraction patterns of the molded chalcogenide-glass lens were evaluated to verify the compositional and structural stability of the glass material under the given molding conditions.