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Study on hydroxy fatty acid contents changes and physiological responses under abiotic stresses in transgenic Camelina

  • Kim, Hyun-Sung;Lee, Hyun-Sook;Lim, Hyun-Gyu;Park, Won;Kim, Hyun-Uk;Lee, Kyeong-Ryeol;Ahn, Sung-Ju
    • Proceedings of the Korean Society of Crop Science Conference
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    • 2017.06a
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    • pp.191-191
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    • 2017
  • Hydroxy fatty acid (HFA) is an important industrial resource that known to be extracted from seeds of Castor or Lesquerella. However, mass production of HFA from those crops are difficult because of their behavior or life cycle. In this study, we applied HFA synthesis related gene FAH12, RcPDAT1, RcLPCAT, RcDGAT2, and RcPDCT on bioenergy crop Camelina sativa. Furthermore, we determined NaCl or cold stress tolerance changes of transgenic Camelina. RcFAH12, RcPDAT1, RcLPCAT, RcDGAT2, and RcPDCT genes were cloned into multigene expression vector which is engineered with seed specific promoter of FAE1 or Napin. Combination of HFA genes multi-expression vector constructs were divided into Set3 (RcFAH12, RcPDAT1-2, RcLPCAT), Set4 (RcFAH12, RcDGAT2, RCPDAT1-2, RcLPCAT), and Set5 (RcFAH12, RcDGAT2, RCPDAT1-2, RcLPCAT, RcPDCT). Transgenic HFA synthesis Camelina plants were generated using agrobacterium-mediated vacuum infiltration system. Results of fatty acid composition of T1 transgenic Camelina seeds analyzed by GC-MS showed 9.5, 9.0, and 13.6 % of HFA contents in Set3#6, Set4#8, and Set5#10, respectively. Therefore, seeds of T2 generation were harvest from Set5#10 which is shown highest HFA contents, and, 17.7, 8.1 and 10.5 % of HFA contents were determined in Set5#10-5, Set5#10-8, and Set#10-10, respectively. However, 7.7% of C18:2 and 22.3 % of C18:3 among unsaturated fatty acids were decreased in Set5#10-5 than WT. Meanwhile, we confirmed abiotic stress responses in T2 transgenic Camelina Set5#10-5 and Set5#10-10 under 0, 100, 150, and 200 mM NaCl or 25, 15, and $10^{\circ}C$ temperature for 5 weeks. Both Set5#10-5 and Set5#10-10 showed lower growth in height than WT in control and NaCl condition. Growth of leaf length and width were similar in WT and Set5#10-10 but lower in Set5#10-5 under NaCl stress. Number of opened flowers showed that both transgenic Camelina were lower than WT under normal condition. But, WT and Set5#10-10 showed similar opened flower number in 100 and 200 mM NaCl. In cold stress, 15 and $10^{\circ}C$ treatment for 5 weeks did not showed significant changes in between WT and both transgenic lines even they showed different growth rate in control condition. Taken together, growth and development are delayed by expression of exogenous HFA related genes in transgenic lines but relative abiotic stress sensitivity is similar with WT. In conclusion, reduced C18:2 or C18:3 fatty acid composition of seed by HFA synthesis is resulted from lack of resource supplement for development at seedling stage but it is not affect NaCl and cold stress tolerance.

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Production of Poly-3-hydroxybutyrate from Xylose by Bacillus megaterium J-65 (Bacillus megaterium J-65에 의한 xylose로부터 poly-3-hydroxybutyrate 생산)

  • Jun, Hong-Ki;Jin, Young-Hi;Kim, Hae-Nam;Kim, Yun-Tae;Kim, Sam-Woong;Baik, Hyung-Suk
    • Journal of Life Science
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    • v.18 no.12
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    • pp.1625-1630
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    • 2008
  • A microorganism capable of producing high level of poly-3-hydoxybutyrate (PHB) from xylose was isolated from soil. The isolated strain J-65 was identified as Bacillus megaterium based on the morphological, biochemical and molecular biological characteristics. The optimum temperature and pH for the growth of B. megaterium J-65 were $37^{\circ}C$ and 8.0, respectively. The optimum medium composition for the cell growth was 2% xylose, 0.25% $(NH_4)_2SO_4$, 0.3% $Na_2HPO_4{\cdot}12H_2O$, and 0.1% $KH_2PO_4$. The optimum condition for PHB accumulation was same to the optimum condition for cell growth. Copolymer of ${\beta}$-hydroxybutyric and ${\beta}$-hydroxyvaleric acid was produced when propionic acid was added to shake flasks containing 20 g/l of xylose. Fermenter culture was carried out to produce the high concentration of PHB. In batch culture, cell mass was 9.82 g/l and PHB content was 35% of dry cell weight. PHB produced by B. megaterium J-65 was identified as homopolymer of 3-hydoxybutyric acid by GC and NMR.

Antifungal activity of extracts from Chamaecyparis obtusa and Pseudotsuga menziesii against Trichoderma spp. (국내산 침엽수 추출물의 Trichoderma spp.에 대한 항균활성)

  • Jung, Ji-Young;Kim, Ji-Woon;Kim, Yeong-Suk;Park, Han-Min;Lee, Byung-Hyun;Choi, Myung-Suk;Yang, Jae-Kyung
    • Journal of agriculture & life science
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    • v.45 no.4
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    • pp.1-11
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    • 2011
  • The aim of this study was development of natural antifungal compounds from softwood. We investigated antifungal activities of extracts from Pseudotsuga menziesii and Chamaecyparis obtusa against Tricholderma genus which is virus causing green mold disease and analyzed antifungal compounds by Gas chromatography -Mass Spetrometer. Extracts from P. menziesii had inhibition activities against Tricholderma genus on 1,000 ppm and had high antifungal activities against T. viride by 70.1%, T. harzianum by 67.3% and T. aggressivum by 64.7% on 4,000 ppm. And extracts from C. obtusa had antifungal activities against Tricholderma genus on 1,000 ppm and had high antifungal activities against T. viride by 63.2%, T. harzianum by 59.3% and T. aggressivum by 59.1% on 4,000 ppm. But mixing compounds which are made from P. menziesii and C. obtusa extracts by variety ratio had lower antifungal activities than original extracts. Main antifungal active components of P. menziesii extracts against Tricholderma genus were 2-Isopropoxy-ethylamine 46.5%, epifluorohydrin 8.6%, trans-2,3-Di-methyloxirane 7.6%, (IR)-(-)-Myrtenal 6.0%, 2-Methoxy-4-Vinylphenol 3.9% and benzaldehyde 2.8%. In case of C. obtusa extracts, they were ${\alpha}$-Terpinenyl acetate 14.9%, Sabinene 10.9%, dl-Limonene 9.6%, ${\alpha}$-Terpinolene 7.5% and ${\alpha}$-Pinene 7.1%. As mentioned above, these results revealed extracts from P. menziesii and C. obtusa of softwood could be used as potential agents to inhibit Trichoderma genus.

Comparative Study on the Composition of Floral Volatile Components in the Flowering Stages of Robinia pseudoacacia L. (아까시나무(Robinia pseudoacacia L.) 꽃의 개화 단계별 향기성분 조성 비교)

  • Jung, Je Won;Lee, Hyun Sook;Noh, Gwang Rae;Lee, Andosung;Kim, Moon Sup;Kim, Sea Hyun;Kwon, Hyung Wook
    • Journal of Apiculture
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    • v.32 no.3
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    • pp.139-146
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    • 2017
  • Floral scent emitted from many plants is the critical factors for pollinator attraction and defense for adaptation in environments. The fragrance components of flowers are different in composition by geographical origins, climate factors and the development stages of flowers. In the present study, we investigated the volatile-floral compounds in flowers of Robinia pseudoacacia L. and defined the chemical contribution for flowering periods. The volatile compounds analysis was performed by gas chromatography with mass selective detector after solid phase microextraction (SPME). We reported different compositional features of fragrance compounds according to flowering periods. The abundant compounds identified in stage 1 were ${\alpha}$-pinene (66.80%) and ${\beta}$-pinene (26.53%). Those of the stage 2 were (Z)-${\beta}$-ocimene (37.57%), ${\alpha}$-pinene (15.16%), benzaldehyde (16.63%), linalool (12.13%). The volatiles of stage 3 comprised an abundance of (Z)-${\beta}$-ocimene (64.94%), ${\alpha}$-pinene (9.84%), linalool (8.92%), benzaldehyde (1.71%). Leaf volatiles were distinct from those in the reproductive plant parts by their high relative amount of (E)-${\beta}$-ocimene (23.50%) and (Z)-3-Hexenyl acetate (27.87%). Differences in flower scents of the different stages and leaves are discussed in light of biochemical constraints on volatile chemical synthesis and of the role of flower scent in evolutionary ecology of R. pseudoacacia.

A study on the calibration characteristics of organic fatty acids designated as new offensive odorants by cryogenic trapping-thermal desorption technique (유기지방산 신규악취물질에 대한 저온농축 열탈착방식 (Thermal desorber)의 검량특성 연구)

  • Ahn, Ji-Won;Kim, Ki-Hyun;Im, Moon-Soon;Ju, Do-Weon
    • Analytical Science and Technology
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    • v.22 no.6
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    • pp.488-497
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    • 2009
  • In this study, analytical methodology for several organic fatty acids (OFA: propionic acid (PA), butyric acid (BA), isovaleric acid (IA), and valeric acid (VA)) designated as new offensive odorants in Korea (as of year 2010) was investigated along with some odorous VOCs (styrene, toluene, xylene, methyl ethyl ketone, methyl isobutyl ketone, butyl acetate, and isobutyl alcohol). For this purpose, working standards (WS) containing all of these 13 compounds were loaded into adsorption tube filled with Tenax TA, and analyzed by gas chromatography (GC) system thermal desorber interfaced with. The analytical sensitivities of organic fatty acids expressed in terms of detection limit (both in absolute mass (ng) and concentration (ppb)) were lower by 1.5-2 times than other compounds (PA: 0.24 ng (0.16 ppb), BA: 0.19 ng (0.11 ppb), IA: 0.15 ng (0.07 ppb), and VA: 0.28 ng (0.13 ppb)). The precision of BA, IA, and VA, if assessed in terms of relative standard error (RSE), maintained above 5%, while the precison of other compounds were below 5%. The reproducibility of analysis improved with the aid of internal standard calibration (PA: $1.1{\pm}0.4%$, BA: $10{\pm}0.46$, IA; $12{\pm}0.3%$, VA: $4{\pm}0.1%$), respectively. The results of this study showed that organic fatty acid can be analyzed using adsorption tube and thermal desorber in a more reliable way to replace alkali absorption method introduced in the odor prevention law of the Korea Ministry of Environment (KMOE).

Radiosynthesis of $[^{11}C]6-OH-BTA-1$ in Different Media and Confirmation of Reaction By-products. ($[^{11}C]6-OH-BTA-1$ 조제 시 생성되는 부산물 규명과 반응용매에 따른 표지 효율 비교)

  • Lee, Hak-Jeong;Jeong, Jae-Min;Lee, Yun-Sang;Kim, Hyung-Woo;Lee, Eun-Kyoung;Lee, Dong-Soo;Chung, June-Key;Lee, Myung-Chul
    • Nuclear Medicine and Molecular Imaging
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    • v.41 no.3
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    • pp.241-246
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    • 2007
  • Purpose: $[^{11}C]6-OH-BTA-1$ ([N-methyl-$^{11}C$]2-(4'-methylaminophenyl)-6-hydroxybenzothiazole, 1), a -amyloid imaging agent for the diagnosis of Alzheimer's disease in PET, can be labeled with higher yield by a simple loop method. During the synthesis of $[^{11}C]1$, we found the formation of by-products in various solvents, e.g., methylethylketone (MEK), cyclohexanone (CHO), diethylketone (DEK), and dimethylformamide (DMF). Materials and Methods: In Automated radiosynthesis module, 1 mg of 4-aminophenyl-6-hydroxybenzothiazole (4) in 100 l of each solvent was reacted with $[^{11}C]methyl$ triflate in HPLC loop at room temperature (RT). The reaction mixture was separated by semi-preparative HPLC. Aliquots eluted at 14.4, 16.3 and 17.6 min were collected and analyzed by analytical HPLC and LC/MS spectrometer. Results: The labeling efficiencies of $[^{11}C]1$ were $86.0{\pm}5.5%$, $59.7{\pm}2.4%$, $29.9{\pm}1.8%$, and $7.6{\pm}0.5%$ in MEK, CHO, DEK and DMF, respectively. The LC/MS spectra of three products eluted at 14.4, 16.3 and 17.6 mins showed m/z peaks at 257.3 (M+1), 257.3 (M+1) and 271.3 (M+1), respectively, indicating their structures as 1, 2-(4'-aminophenyl)-6-methoxybenzothiazole (2) and by-product (3), respectively. Ratios of labeling efficiencies for the three products $([^{11}C]1:[^{11}C]2:[^{11}C]3)$ were $86.0{\pm}5.5%:5.0{\pm}3.4%:1.5{\pm}1.3%$ in MEK, $59.7{\pm}2.4%:4.7{\pm}3.2%:1.3{\pm}0.5%$ in CHO, $9.9{\pm}1.8%:2.0{\pm}0.7%:0.3{\pm}0.1%$ in DEK and $7.6{\pm}0.5%:0.0%:0.0%$ in DMF, respectively. Conclusion: The labeling efficiency of $[^{11}C]1$ was the highest when MEK was used as a reaction solvent. As results of mass spectrometry, 1 and 2 were conformed. 3 was presumed.