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Relationship between porcine miR-20a and its putative target low-density lipoprotein receptor based on dual luciferase reporter gene assays

  • Ding, Yueyun;Zhu, Shujiao;Wu, Chaodong;Qian, Li;Li, DengTao;Wang, Li;Wan, Yuanlang;Zhang, Wei;Yang, Min;Ding, Jian;Wu, Xudong;Zhang, Xiaodong;Gao, Yafei;Yin, Zongjun
    • Asian-Australasian Journal of Animal Sciences
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    • v.32 no.7
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    • pp.922-929
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    • 2019
  • Objective: Mutations in low-density lipoprotein receptor (LDLR), which encodes a critical protein for cholesterol homeostasis and lipid metabolism in mammals, are involved in cardiometabolic diseases, such as familial hypercholesterolemia in pigs. Whereas microRNAs (miRNAs) can control LDLR regulation, their involvement in circulating cholesterol and lipid levels with respect to cardiometabolic diseases in pigs is unclear. We aimed to identify and analyze LDLR as a potential target gene of SSC-miR-20a. Methods: Bioinformatic analysis predicted that porcine LDLR is a target of SSC-miR-20a. Wild-type and mutant LDLR 3'-untranslated region (UTR) fragments were generated by polymerase chain reaction (PCR) and cloned into the pGL3-Control vector to construct pGL3 Control LDLR wild-3'-UTR and pGL3 Control LDLR mutant-3'-UTR recombinant plasmids, respectively. An miR-20a expression plasmid was constructed by inserting the porcine premiR-20a-coding sequence between the HindIII and BamHI sites in pMR-mCherry, and constructs were confirmed by sequencing. HEK293T cells were co-transfected with the miR-20a expression or pMR-mCherry control plasmids and constructs harboring the corresponding 3'-UTR, and relative luciferase activity was determined. The relative expression levels of miR-20a and LDLR mRNA and their correlation in terms of expression levels in porcine liver tissue were analyzed using reverse-transcription quantitative PCR. Results: Gel electrophoresis and sequencing showed that target gene fragments were successfully cloned, and the three recombinant vectors were successfully constructed. Compared to pMR-mCherry, the miR-20a expression vector significantly inhibited wild-type LDLR3'-UTR-driven (p<0.01), but not mutant LDLR-3'-UTR-driven (p>0.05), luciferase reporter activity. Further, miR-20a and LDLR were expressed at relatively high levels in porcine liver tissues. Pearson correlation analysis revealed that porcine liver miR-20a and LDLR levels were significantly negatively correlated (r = -0.656, p<0.05). Conclusion: LDLR is a potential target of miR-20a, which might directly bind the LDLR 3'-UTR to post-transcriptionally inhibit expression. These results have implications in understanding the pathogenesis and progression of porcine cardiovascular diseases.

Comparison of Post-Harvest Character and Storability at Several Temperature for Lactuca indica L. Baby and Adult Leaves (왕고들빼기 어린잎과 성체의 수확 후 특성과 온도별 저장성 비교)

  • Kim, Ju Young;Han, Su Jeong;Wang, Lixia;Choi, In-Lee;Kang, Ho-Min
    • Journal of Bio-Environment Control
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    • v.28 no.2
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    • pp.172-177
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    • 2019
  • Lactuca indica L. ('Seonhyang') baby leaves were harvested after cultivation for 4 weeks (less than 10 cm plant height) and adults were cultivated for 8 weeks ($20{\pm}5cm$ plant height). The respiration rate and the ethylene production rate of Lactuca indica leaves were higher than those of the baby leaves but the DPPH radical scavenging ability was lower than baby leaves. The $L^*$, $a^*$ and $b^*$ values did not show any significant difference between baby leaves and adult leaves but the chlorophyll content was higher in adult leaves. All adult and baby leaves of Lactuca indica were stored at $2^{\circ}C$, $8^{\circ}C$, and $20^{\circ}C$, respectively. The higher the storage temperature, the higher the fresh weight loss rate and weight loss rate of adult leaves was lower at $2^{\circ}C$ and $8^{\circ}C$. The visual quality of Lactuca indica leaves were determined by the panel test during storage and it deteriorated faster as the storage temperature increased. The shelf life that calculated the period of maintaining higher than 3 points of visual quality was longer than 1.6 days at $2^{\circ}C$, 1.4 days at $8^{\circ}C$ and 1.5 days at $20^{\circ}C$. The oxygen and carbon dioxide concentrations within the package of Lactuca indica leaves were similar to those in atmosphere. The chlorophyll content was maintained higher at lower storage temperature in the last storage day and the off-odor was higher in baby leaves than in the adult leaves of Lactuca indica L.

Changes in Marketability of Strawberry 'Maehyang' for Export as Affected by Concentration of Gaseous Chlorine Dioxide Treatment (이산화염소 가스 처리 농도에 따른 수출 딸기 '매향'의 상품성 변화)

  • Kim, Hye Min;Hwang, Seung Jae
    • Journal of Bio-Environment Control
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    • v.28 no.2
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    • pp.166-171
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    • 2019
  • This study was conducted to investigate the effect of gaseous chlorine dioxide ($ClO_2$) concentration and treatment method for maintaining marketability in strawberries 'Maehyang' for export. After harvesting strawberry colored with $60{\pm}5%$ of the skin, and the gaseous $ClO_2$ was applied as four treatments in the cold store set with $10^{\circ}C$ which were as follows: i) non-treatment (Control), ii) $0.2mg{\cdot}L^{-1}$ for 30 minutes, iii) $0.4mg{\cdot}L^{-1}$ for 30 minutes, and iv) continuously exposed at $0.4mg{\cdot}L^{-1}$ during the storage period. Weight loss, firmness, soluble solids content, color, incidence of gray mold, and quality grade of strawberries were investigated every 3 days during 16 days storage in cold store. The weight loss was consistently high at $0.2mg{\cdot}L^{-1}\;ClO_2$ treatment, and the weight loss was lower than other treatments when gaseous $ClO_2$ was continuously treated. Firmness was significantly higher at 0.2 and $0.4mg{\cdot}L^{-1}$ $ClO_2$ treatment on the 13th day of storage. Soluble solids content tended to below in continuous gaseous $ClO_2$ treatment. The colors showed no tendency as affected by concentrations and treatment methods of gaseous $ClO_2$. No gray mold was observed in the continuous gaseous $ClO_2$ treatment until the 13th day of storage. The quality grade was lower in gaseous $ClO_2$ treatment group than in the control. As a result, it was possible to decrease the weight loss and the incidence of gray mold by continuously treating the gaseous $ClO_2$ to strawberry 'Maehyang' for export during the storage period after harvest. However, further research is needed on method to maintain quality grades of fruits.

Epidemiology of Staphylococcus aureus Bacteremia in Children at a Single Center from 2002 to 2016 (단일 기관에서 2002년부터 2016년까지 발생한 소아 황색포도알균 균혈증의 역학)

  • Lim, Seonhee;Ha, Seok Gyun;Tchah, Hann;Jeon, In Sang;Ryoo, Eell;Son, Dong Woo;Cho, Hye Jung;Sun, Yong Han;Kim, Hyo Jung;Ahn, Jung Min;Cho, Hye-Kyung
    • Pediatric Infection and Vaccine
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    • v.26 no.1
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    • pp.11-21
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    • 2019
  • Purpose: We aimed to investigate the epidemiological characteristics of Staphylococcus aureus bacteremia in Korean children. Methods: We retrospectively collected and analyzed data from the medical records of the patients with S. aureus bacteremia ${\leq}18years$ of age in Gil Medical Center from 2002 to 2016. Results: A total of 212 SAB cases were detected. The annual incidence of SAB from 2002 to 2016 ranged from 0.77 to 1.95 per 1,000 patients hospitalized. The neonate group (<28 days of age) and the pediatric group (28-18 years of age) were 51.4% (n=109) and 48.6% (n=103), respectively. According to the origin of infection, there were 93 cases (43.9%) of community-associated (CA)-SAB and 119 cases (56.1%) of healthcare-associated (HA)-SAB. The rates of HA-SAB among the neonate group and among the pediatric group were 64.2% and 47.6%, respectively (P=0.015). There was no difference in complications between CA-SAB and HA-SAB, but mortality was higher in HA-SAB. The proportion of methicillin-resistance S. aureus (MRSA) was the highest in neonates (88.1%), decreased with age, and was 36.4%-37.5% among children aged ${\geq}5years$. The MRSA proportion was 72.2%, showing no consistent trend over the period. Conclusions: The annual incidence of SAB and the proportion of MRSA in SAB remained constant in the recent 15 years in children. Judicious decision of antimicrobial agents for treatment considering the patient's age and the origin of infection is necessary.

Dissipation of Bifenthrin and Chlorothalonil in Crown Daisy during Cultivation and their Biological Half-lives (쑥갓 중 bifenthrin과 chlorothalonil의 경시적 잔류량 변화 및 생물학적 반감기)

  • Kang, Min-Seong;Park, Po-Hyun;Kim, Ki-Yu;Lim, Bu-Geon;Ryu, Kyong-Shin;Lee, You-Jin;Lim, Jeong-Hwa;Kang, Choong-Won;Kim, Youn-Ho;Lee, Sun-Young;Seo, Jeong-Hwa;Park, Yong-Bae;Yoon, Mi-Hye
    • Journal of Food Hygiene and Safety
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    • v.34 no.2
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    • pp.191-198
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    • 2019
  • This study investigated the residual characteristics of bifenthrin and chlorothalonil in crown daisy and suggested pre-harvest residue limits (PHRLs) based on their dissipation patterns and biological half-lives. The samples for residue analysis were harvested at 0 (3 hr), 1, 3, 5, 7, 9, 11, 13, 15, 18, 22 and 26 days after treatment, and analyzed by $GC/{\mu}-ECD$ and TOF/MS. The limit of quantitation (LOQs) of bifenthrin and chlorothalonil were 0.0046 mg/kg and 0.0007 mg/kg, respectively. Recoveries ranged from $88.67{\pm}7.97%$ and $99.90{\pm}16.03%$, showing that this method is appropriate for the analysis of the pesticide residues in crown daisy. Being well within first order kinetics, the biological half-lives of the pesticide residues in crown daisy were 9.63 days for bifenthrin and 6.54 days for chlorothalonil. The PHRLs of bifenthrin and chlorothalonil were recommended as 11.70 mg/kg and 24.10 mg/kg for 26 days before harvest, respectively.

Development and Validation of an Analytical Method for Fenpropimorph in Agricultural Products Using QuEChERS and LC-MS/MS (QuEChERS법과 LC-MS/MS를 이용한 농산물 중 Fenpropimorph 시험법 개발 및 검증)

  • Lee, Han Sol;Do, Jung-Ah;Park, Ji-Su;Cho, Sung Min;Shin, Hye-Sun;Jang, Dong Eun;Choi, Young-Nae;Jung, Yong-hyun;Lee, Kangbong
    • Journal of Food Hygiene and Safety
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    • v.34 no.2
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    • pp.115-123
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    • 2019
  • An analytical method was developed for the determination of fenpropimorph, a morpholine fungicide, in hulled rice, potato, soybean, mandarin and green pepper using QuEChERS (Quick, Easy, Cheap, Effective, Rugged and Safe) sample preparation and LC-MS/MS (liquid chromatography-tandem mass spectrometry). The QuEChERS extraction was performed with acetonitrile followed by addition of anhydrous magnesium sulfate and sodium chloride. After centrifugation, d-SPE (dispersive solid phase extraction) cleanup was conducted using anhydrous magnesium sulfate, primary secondary amine sorbents and graphitized carbon black. The matrix-matched calibration curves were constructed using seven concentration levels, from 0.0025 to 0.25 mg/kg, and their correlation coefficient ($R^2$) of five agricultural products were higher than 0.9899. The limits of detection (LOD) and quantification (LOQ) were 0.001 and 0.0025 mg/kg, respectively, and the limits of quantification for the analytical method were 0.01 mg/kg. Average recoveries spiked at three levels (LOQ, $LOQ{\times}10$, $LOQ{\times}50$, n=5) and were in the range of 90.9~110.5% with associated relative standard deviation values less than 5.7%. As a result of the inter-laboratory validation, the average recoveries between the two laboratories were 88.6~101.4% and the coefficient of variation was also below 15%. All optimized results were satisfied the criteria ranges requested in the Codex guidelines and Food Safety Evaluation Department guidelines. This study could serve as a reference for safety management relative to fenpropimorph residues in imported and domestic agricultural products.

Prediction of Propylene/Propane Separation Behavior of Na-type Faujasite Zeolite Membrane by Using Gravimetric Adsorption (중량식흡착 거동에 기초한 Na형 Faujasite 제올라이트 분리막의 프로필렌/프로페인 분리 거동 예측 연구)

  • Hwang, Juyeon;Min, Hae-Hyun;Park, You-In;Chang, Jong-San;Park, Yong-Ki;Cho, Churl-Hee;Han, Moon-Hee
    • Membrane Journal
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    • v.28 no.6
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    • pp.432-443
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    • 2018
  • In this study, propylene/propane separation behavior of Na-type faujasite zeolite membranes is predicted by observing gravimetric adsorptions of propylene and propane on zeolite 13X. The gravimetric adsorptions were measured by using a magnetic suspension balance (MSB) at temperatures of 323, 343, 363 K and a pressure range of 0.02-1 bar. The pressure was increased at 0.1 bar intervals. As adsorption temperature increased, adsorptions of propylene and propane decreased and propylene/propane adsorption selectivity increased. Also, the diffusion coefficients of propylene and propane were increased as the adsorption temperature increased, following the Arrhenius equation. The maximum propylene/propane diffusion selectivity was 0.9753 at 323 K. The perm-selectivity was calculated from the adsorption data of zeolite 13X and compared with the perm-selectivity measured in the single gas permeation experiment for the Na-type faujasite zeolite membrane. The maximum values for the calculated and measured perm-selectivities were observed at a temperature of 323 K. It could be concluded that the prediction of propylene/propane separation of surface diffusion-based membrane by using gravimetric adsorption data is reasonable. Therefore, it is expected that this prediction method can be applied to the screening of adsorption-based microporous membrane for propylene/propane separation.

Introduction of the International Standardization of ISO in the Production and Quality of Herbal Medicines and a Review of Countermeasures (한약재 생산 및 품질부문의 ISO국제표준화 등재현황 소개 및 대응방안 고찰)

  • Kim, Yong Il;Kang, Young Min;Han, Sin-Hee;Hur, Mok;Kim, Young Guk;Chang, Jae Ki
    • Journal of the Korean Society of International Agriculture
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    • v.30 no.4
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    • pp.257-268
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    • 2018
  • Recently, the international standardization of ISO in the field of Oriental Herbal Medicine has been progressing rapidly under the direction of China's promotion. China's intention to promote international standardization is to extend its influence to the world and beyond the domestic market. The Oriental medicine system in East Asia has similar roots in academic terms, but the medicines that can be supplied and received in each country are different and have developed independently. The international standardization of medicinal herbs is expected to function in a direction that weakens such differentiation and independence. From a commercial point of view, international standardization is no different from creating evaluation criteria for oriental medicinal products, and it is expected that its potential impact on domestic and overseas markets and producers will be large. In particular, the international standardization centered on China can lead to favorable evaluation criteria for China, which may further negatively affect the market competitiveness of domestic raw materials, which have been pushed back by Chinese manufacturers. If the domestic production base is weakened, not only will the farmers suffer but the supply and demand of raw materials will also be manipulated, safety management control will be reduced, and the development of oriental herbal products using domestic raw materials will be hurt. Therefore, in the promotion of international standardization, it is necessary not only to reflect the value of Korean herbal medicine but also to provide strategic responses to protect the domestic production base. However, in the case of recent initiatives, there is no precedent in analyzing influence on the production partners and the related industries. In addition, there are few related papers and reports on the subject, so the publicity process has not been done sufficiently. In response to this, this study will examine the countermeasures against the international standardization of herbal medicines through reviewing its present status and evaluating the agenda of the Korean initiative.

Effect of Semisulcospira libertina Extracts from Different Extraction Processes on Liver Cell Toxicity and Ethanol Metabolism (간세포 독성과 에탄올 대사에서 추출 조건에 따른 다슬기 추출물의 효과)

  • Cho, Kyoung Hwan;Choo, Ho Jin;Seo, Min Gyun;Kim, Jong Cheol;Shin, Yu Jin;Ryu, Gi Hyung;Cho, Hee Young;Jeong, Chi-Young;Hah, Young-Sool
    • Food Engineering Progress
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    • v.21 no.2
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    • pp.158-166
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    • 2017
  • Although Semisulcospira libertina is generally regarded as a supplement for the alleviation of alcohol hangover, little is known about its effects on cell metabolism. Therefore, this study was conducted to analyze the constituents of the extracts prepared using different extraction methods and to compare their biochemical properties. The amino acid contents were found to be much higher in acidic and enzymatic hydrolysates than hot water extracts from S. libertina. DPPH radical scavenging activities in acidic and enzymatic hydrolysates were higher than those of hot water extracts. Three types of S. libertina hydrolysate was added to HepG2 cells damaged by acetaminophen (AAP), after which the survival rate of HepG2 cell were measured. In addition, lactate dehydrogenase (LDH) activities in the culture media were evaluated. The survival rates of HepG2 cells were $77.0{\pm}4.3%$ and $81.5{\pm}1.3%$ at 3 h and 5h enzymatic hydrolysates, respectively. These cell survival rates were higher compared to those of the negative control group ($67.8{\pm}4.3%$) treated only with acetaminophen. Cellular toxicities induced by treatment with AAP were also significantly alleviated in response to treatment with the extracts of S. libertina. In addition, the activities of 2 key enzymes that metabolize ethanol, alcohol dehydrogenase and aldehyde dehydrogenase, were upregulated by 4.7- and 2.7-fold respectively in response to treatment with a 3 h enzymatic hydrolysate of S. libertina. Taken together, these results provide biochemical evidence of the method by which S. libertina exerts its biological functions, including the alleviation of alcohol hangover and the protection of liver cells against toxic insults.

'Hyangcheola', A New Fragrant Rice Variety with High Iron and Zinc Content (철과 아연 함량이 높고 향기가 있는 벼 '향철아')

  • Jeong, Eung-Gi;Ahn, Eok-Keun;Won, Yong-Jae;Lee, Jeong-Heui;Lee, Sang-Bok;Jeong, Jong-Min;Oh, Sea-Kwan;Choi, Yong-Hwan;Lee, Jeom-Ho;Suh, Jung-Pil;Hyun, Ung-Jo;Jung, O-Young;Kim, Bo-Kyeong;Son, Ji-Young
    • Korean Journal of Breeding Science
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    • v.50 no.4
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    • pp.472-477
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    • 2018
  • "Hyangcheola" is a new japonica rice variety developed as a cross between "Joryeong" and "Seolhyangchal" by the rice breeding team at NICS in 2014. The heading date of "Hyangcheola" is July 28 in the middle plain area, which is 8 days earlier than that of "Hwaseong." "Hyangcheola" has a 76 cm long culm, which is 8 cm shorter than "Hwaseong" and has 86 spikelets per panicle. The viviparous germination rate of "Hyangcheola" was 27.9%. It showed resistance to stripe virus but susceptibility to bacterial blight (K1, K2, and K3 races), dwarf and black streak dwarf viruses and planthoppers. The milled rice of this variety exhibits translucent and medium short grain shape. The protein content of "Hyangcheola" was 8.2%, which was 1.5% higher than that of "Hwaseong." "Hyangcheola" is a rice with a fragrant aroma. The content of iron in 100 g of brown "Hyangcheola" rice was 15.12 mg, which is 3.69 mg higher than that of "Hwaseong," and the zinc content was 32.24 mg, which is 8.75 mg more than that of the comparative variety "Hwaseong." The average milled rice yield of "Hyangcheola" was 4.8 MT/ha at our sites under ordinary cultivation (Registration No. 6807).