• Title/Summary/Keyword: molecular analysis

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Development of Species-Specific PCR to Determine the Animal Raw Material (종 특이 프라이머를 이용한 동물성 식품원료의 진위 판별법 개발)

  • Kim, Kyu-Heon;Lee, Ho-Yeon;Kim, Yong-Sang;Kim, Mi-Ra;Jung, Yoo Kyung;Lee, Jae-Hwang;Chang, Hye-Sook;Park, Yong-Chjun;Kim, Sang Yub;Choi, Jang Duck;Jang, Young-Mi
    • Journal of Food Hygiene and Safety
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    • v.29 no.4
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    • pp.347-355
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    • 2014
  • In this study, the detection method was developed using molecular biological technique to distinguish authenticity of animal raw materials. The genes for distinction of species about animals targeted at Cytochrome c oxidase subunit I (COI), Cytochrome b (Cytb), and 16S ribosomal RNA (16S rRNA) genes in mitochondrial DNA. The species-specific primers were designed by that Polymerase Chain Reaction (PCR) product size was around 200 bp for applying to processed products. The target 24 raw materials were 2 species of domestic animals, 6 species of poultry, 2 species of freshwater fishes, 13 species of marine fishes and 1 species of crustaceans. The results of PCR for Rabbit, Fox, Pheasant, Domestic Pigeon, Rufous Turtle Dove, Quail, Tree Sparrow, Barn Swallow, Catfish, Mandarin Fish, Flying Fish, Mallotus villosus, Pacific Herring, Sand Lance, Japanese Anchovy, Small Yellow Croaker, Halibut, Jacopever, Skate Ray, Ray, File Fish, Sea Bass, Sea Urchin, and Lobster raw materials were confirmed 113 bp ~ 218 bp, respectively. Also, non-specific PCR products were not detected in compare species by species-specific primers. The method using primers developed in this study may be applied to distinguish an authenticity of food materials included animal raw materials for various processed products.

Identification of Fatty Acids in the Pulp Oils of Jujube and Their Compsitional Changes in the Ripening Period (대추의 과육지질(果肉脂質)에 존재(存在)하는 지방산(脂肪酸)의 동정(同定)과 숙성(熟成)에 따른 그 조성(組成)의 변화(變化))

  • Woo, Hyo-Kyeng;Kim, Seong-Jin;Park, Sung-Hea;Joh, Yong-Goe
    • Journal of the Korean Applied Science and Technology
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    • v.18 no.1
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    • pp.67-77
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    • 2001
  • In search for several fatty acid with unusual structure in vegetable oils, we have found that unknown peaks were shown on GLC in the analysis of fatty acids of the lipids from the pulp of ripened jujube (Zizypus jujuba var. inermis) fruits. These fatty acids were identified as a series of cis-monoenoic acids with ${\omega}-5$ double bond system such as $C_{14:1{\omega}5}$, $C_{16:1{\omega}5}$ and $C_{18:1{\omega}5}$, including ${\omega}-7$ fatty acid as $C_{16:1{\omega}7}$ and $C_{18:1{\omega}7}$, by GLC, solid-phase extraction silver ion-column chromatographic, GLC-mass spectrometric and IR techniques. First of all, total fatty acid methyl esters were resolved into saturated and branched fatty acid, monoenoic acid, dienoic acid, and trienoic acid fraction, respectively, with 100% dichloromethane (DCM), DCM/acetone (9:1, v/v) 100% acetone, and acetone/ acetonitrile (97:3, v/v) solvent system. Unknown fatty acids were included in the monoenoic fraction and were confirmed to have cis-configuration by IR. Picolinyl esters of monoenoic fatty acids gave distinct molecular ion peak and dominant diagnostic peaks, for example, m/z 317, 220 and 260 fragment for $cis-C_{14:1{\omega}5}$, m/z 345, m/z 248 and 288 fragment for $cis-C_{16:1{\omega}5}$ and m/z 373, m/z 276 and 316 fragment for $cis-C_{18:1{\omega}5}$. In this way the occurrence of $cis-C_{16:1{\omega}7}$ and $cis-C_{18:1{\omega}7}$ could be deduced from the appearance of prominent fragments as m/z 345, 220 and 260, and m/z 373, 248 and 280. Level of total ${\omega}-5$ fatty acids amounted to about 30% in the fatty acid composition with the predominance of $C_{16:1{\omega}5}$ $ (18.7{\sim}25.0%)$, in the semi-ripened and/or ripened samples collected in September 14 ($C_{16:1{\omega}5}$ ; 18.7%, $C_{14:1{\omega}5}$ ; 3.6% and $C_{18:1{\omega}5}$ ; 3.0%), September 22 ($C_{16:1{\omega}5}$ ; 25.0%, $C_{14:1{\omega}5}$ ; 1.4% and $C_{18:1{\omega}5}$ ; 2.6%), and October $7 (C_{16:1{\omega}5}$ ; 24.7%, $C_{14:1{\omega}5}$ ; 7.7% and $C_{18:1{\omega}5}$ ; 2.5%). However, the lipids extracted from unripened jujube in July and August contain these unusual fatty acids as low as negligible. It could be observed that the level of ${\omega}-5$ fatty acids in the pulps increased sharply with an elapse of ripening time of jujube fruits. Other monoenoic fatty acids with ${\omega}-7$ series, $C_{16:1{\omega}7}$ (palmitoleic acid) and $C_{18:1{\omega}7}$ (cis-vaccenic acid) could be detected. And in the lipids of the kernel and leaf of jujube, none of ${\omega}-5$ fatty acids could be detected.

Studies on the Anti-aging Action of Korean Ginseng (I) Comparative Study of Red and White Ginsengs on Anti-aging Action (고려인삼(高麗人蔘)의 노화억제작용(老化抑制作用)에 관한 연구(硏究) 제1보(第1報) 홍삼(紅蔘)과 백삼(白蔘)의 노화억제작용(老化抑制作用) 비교연구(比較硏究))

  • Choi, Jin Ho;Oh, Sung Ki
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.12 no.4
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    • pp.323-335
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    • 1983
  • The inhibitory effects of the extract and crude saponin of red and white ginsengs on lipoperoxide formation in vitro and in vivo were studied and correlated with anti-aging. To this end, antioxidant activity, induction period and lipoperoxide were measured by the methods of EDA, POV and TBA value. And also superoxide dismutase and peroxidase activity were measured by pyrogallol autoxidation method (${\Delta}A$ 420/min) and initial velocity(${\Delta}A$ 436/min), respectively. From HPLC analysis, the PT/PD ratio of red and white ginsengs was found to be 0.561% and 0.401%, respectively, and red ginseng increased the PT/PD ratio in comparison with white ginseng. The EDA activity of red ginseng was higher than that of white ginseng; red ginseng showed stronger antioxidative effect than white ginseng. The inhibitory effect of red ginseng was lower than that of white ginseng during the induction period. It was proved that high molecular coloring substance was deeply related to the initial stage of lipoperoxidation. There was no significant difference between red and white ginsengs in both in vitro and intraperitoneal administration experiments, and red ginseng was more effective than white ginseng in longterm administration. And also inhibitory effect on lipoperoxide formation was mainly occurred in liver, suggesting that the function of liver played an important role in anti-aging actions. From the measurement of superoxide dismutase(SOD) activity for both ginseng groups intraperitoneally and orally administered, it was found that red ginseng group administered extract and crude saponin showed remarkable inhibitory effects in comparison with white ginseng. In particular, orally administered group showed more stronger inhibitory effect on lipid peroxidation in comparison with intraperitoneally administered group. It was also found that the continuous oral administration was more effective than temporary administration. Red ginseng was more notable anti-aging effect in comparison with white ginseng in vivo, and this may be due to the increase of SOD activity in rat-liver. Peroxidase activity also showed similar trend to SOD activity in vitro and in vivo experiments. Red ginseng was not only superior to white ginseng for preservation but also for biochemical and pharmaceutical efficacy.

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Validation of Stem-loop RT-qPCR Method on the Pharmacokinetic Analysis of siRNA Therapeutics (Stem-loop RT-qPCR 분석법을 이용한 siRNA 치료제의 생체시료 분석법 검증 및 약물 동태학적 분석)

  • Kim, Hye Jeong;Kim, Taek Min;Kim, Hong Joong;Jung, Hun Soon;Lee, Seung Ho
    • Journal of Life Science
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    • v.29 no.6
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    • pp.653-661
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    • 2019
  • The first small interfering RNA (siRNA) therapeutics have recently been approved by the Food and Drug Administration in the U.S., and the demand for a new RNA therapeutics bioanalysis method-which is essential for pharmacokinetics, including the absorption, distribution, metabolism, and excretion of siRNA therapeutics-is rapidly increasing. The stem-loop real-time qPCR (RT-qPCR) assay is a useful molecular technique for the identification and quantification of small RNA (e.g., micro RNA and siRNA) and can be applied for the bioanalysis of siRNA therapeutics. When the anti-HPV E6/E7 siRNA therapeutic was used in preclinical trials, the established stem-loop RT-qPCR assay was validated. The limit of detection was sensitive up to 10 fM and the lower limit of quantification up to 100 fM. In fact, the reliability of the established test method was further validated in three intra assays. Here, the correlation coefficient of $R^2$>0.99, the slope of -3.10 ~ -3.40, and the recovery rate within ${\pm}20%$ of the siRNA standard curve confirm its excellent robustness. Finally, the circulation profiles of siRNAs were demonstrated in rat serum, and the pharmacokinetic properties of the anti-HPV E6/E7 siRNA therapeutic were characterized using a stem-loop RT-qPCR assay. Therefore, the stemloop RT-qPCR assay enables accurate, precise, and sensitive siRNA duplex quantification and is suitable for the quantification of small RNA therapeutics using small volumes of biological samples.

Association Study of Zygote Arrest 1 on Semen Kinematic Characteristics in Duroc Boars (두록 정자 운동학적 특성과 Zygote arrest 1 유전자 변이와의 연관성 분석)

  • Lee, Mi Jin;Ko, Jun Ho;Kim, Yong Min;Choi, Tae Jeong;Cho, Kyu Ho;Kim, Young Sin;Jin, Dong Il;Kim, Nam Hyung;Cho, Eun Seok
    • ANNALS OF ANIMAL RESOURCE SCIENCES
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    • v.29 no.4
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    • pp.150-157
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    • 2018
  • The Zygote arrest 1 (ZAR1) gene is known to affect early embryonic development in various vertebrates. In this study, we performed the association analysis to check whether there is any significant relationship between semen kinematic characteristics and the ZAR1 gene. To determine semen kinematic characteristics, we measured motility (MOT), straight-line velocity (VSL), curvilinear velocity (VCL), average path velocity (VAP), linearity (LIN), straightness (STR), amplitude of lateral head displacement (ALH), and beat cross frequency (BCF) of spermatozoa in boars. In order to detect single nucleotide polymorphisms (SNPs), we extracted genomic DNA from multiple Duroc boars, and then subsequently used them in sequencing reactions. As a result, three SNPs were detected in the intronic region of ZAR1 gene (g.2435T>C in intron 2, g.2605G>A and g.4633A>C in intron 3 ). SNPs g.2435T>C and g.2605G>A were significantly associated with MOT (p<0.01) and VSL (p<0.05), and g.4633A

Cementless Total Hip Arthroplasty Using Ceramic Femoral Head on Cross-Linked Ultra-High-Molecular Weight Polyethylene Liner in Patients Older than 65 Years: Minimum Five-Year Follow-Up Results (세라믹 대퇴 골두 및 교차결합 초고분자량 폴리에틸렌 라이너를 이용한 65세 이상 무시멘트형 인공 고관절 전치환술: 최소 5년 중기 추시 결과)

  • Yun, Ho Hyun;Cheong, Ji Young;Sim, Hyun Bo;Park, Jae Hong
    • Journal of the Korean Orthopaedic Association
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    • v.53 no.6
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    • pp.490-497
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    • 2018
  • Purpose: To evaluate the utility of ceramic-on-polyethylene articular bearing surface when cementless total hip arthroplasty is performed in patients older than 65 years through an analysis of the minimum five-year follow-up results using the ceramic femoral head and cross-linked polyethylene liner. Materials and Methods: From March 2010 to September 2012, 51 patients (56 hips) who were older than 65 years were enrolled in this retrospective study. The mean age at surgery was $70.9{\pm}5.1years$ old. A clinical assessment was analyzed using the Harris hip score. For the radiographic assessment, the cup inclination and anteversion, stem alignment, and wear amount were measured. The postoperative complications were also determined. Results: The mean Harris hip score was improved from preoperative 48 points to postoperative 87 points (p<0.05). The mean cup inclination was $40.9^{\circ}{\pm}6.4^{\circ}$ and the mean cup anteversion was $20.3^{\circ}{\pm}8.1^{\circ}$. The mean cup anteversion of the elevated liner-used group (16 cases) was $14.3^{\circ}{\pm}7.9^{\circ}$ and the mean cup anteversion of the neutral liner used group (40 cases) was $22.4^{\circ}{\pm}9.1^{\circ}$ (p<0.05). The mean stem alignment angle was $0^{\circ}$ (range, varus $4^{\circ}$-valgus $4^{\circ}$). The mean linear wear amount was $0.458{\pm}0.041mm$ and the average annual linear wear rate was $0.079{\pm}0.032mm/yr$. Six cases (10.7%) of intraoperative periprosthetic femoral fractures were encountered. Conclusion: Based on these results, the use of a ceramic-on-polyethylene articular bearing surface in elderly patients with cementless total hip arthroplasty is beneficial. On the other hand, careful effort is needed to prevent intraoperative periprosthetic femoral fractures.

Proteome analysis of storage roots of two sweet potato cultivars with contrasting low temperature tolerance during storage (저온 저장 감수성 및 저항성 고구마 품종에서 저온 반응성 단백질체 연구)

  • Kim, Yun-Hee;Ji, Chang Yoon;Kim, Ho Soo;Chung, Jung-Sung;Choi, Sung Hwan;Kwak, Sang-Soo;Lee, Jeung Joo
    • Journal of Plant Biotechnology
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    • v.49 no.2
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    • pp.118-123
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    • 2022
  • To obtain information on the molecular mechanism underlying the low temperature tolerance of sweet potato [Ipomoea batatas (L.) Lam], the proteome expressed in the sweet potato cultivar Xushu 15-1 with high cold storage tolerance and in the cultivar Xushu 15-4 with low cold storage tolerance was analyzed using 2-D and MALDI-TOF/TOF analyses. Compared with the control (without cold treatment), four protein spots were newly expressed in Xushu 15-1. The expression level of one protein spot was higher in Xushu 15-4 than in Xushu 15-1. Spot 2, which was newly expressed in Xushu 15-1, was identified as sporamin. Assessment of the change in protein expression levels over 8 weeks in the storage roots of the two cultivars treated at 4℃ revealed no significant difference in the expression levels in Xushu 15-1 over time. However, in Xushu 15-4, the expression level of one protein spot increased, while those of four spots decreased. Of the proteins with reduced expression levels, spots 7 and 8 were identified as actin and spots 9 and 10 were identified as fructokinase-like proteins. The present results are expected to enhance the understanding of the complex mechanism underlying the low temperature tolerance of sweet potatoes during storage and can be used to identify candidate genes for the development of new varieties of sweet potatoes with improved low temperature tolerance during cold storage in the future.

Production of Poly (3-Hydroxybutyrate-co-3-Hydroxyvalerate) by Bacillus sp. EMK-5020 Using Makgeolli Lees Enzymatic Hydrolysate and Propionic Acid as Carbon Sources (막걸리 주박 가수분해 산물과 propionic acid를 탄소원으로 이용한 Bacillus sp. EML-5020 균주로부터 poly (3-hydroxybutyrate-co-3-hydroxyvalerate) 생합성)

  • Kwon, Kyungjin;Kim, Jong-Sik;Chung, Chung-Wook
    • Journal of Life Science
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    • v.32 no.7
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    • pp.510-522
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    • 2022
  • In this study, to biosynthesize PHA with properties more similar to polypropylene, a Bacillus sp. EMK-5020 strain that biosynthesized poly (3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) was isolated from soil. Bacillus sp. EMK-5020 strain biosynthesized PHBV containing 1.3% 3-hydroxyvalerate (3HV) using reducing sugar contained in Makgeolli lees enzymatic hydrolysate (MLEH) as a single carbon source. As the amount of propionic acid, which was added as a second carbon source, increased, the content of 3HV also increased. PHBV containing up to 48.6% of 3HV was synthesized when 1.0 g/l of propionic acid was added. Based on these results, the strain was cultured for 72 hr in a 3 l fermenter using reducing sugar in MLEH (20 g/l) and propionic acid (1 g/l) as the main and secondary carbon sources, respectively. As a result, 6.4 g/l DCW and 50 wt% of PHBV (MLEH-PHBV) containing 8.9% 3HV were biosynthesized. Through gel permeation chromatography and thermogravimetric analysis, it was confirmed that the average molecular weight and the decomposition temperature of MLEH-PHBV were 152 kDa and 273℃, respectively. In conclusion, the Bacillus sp. EMK-5020 strain could biosynthesize PHBV containing various 3HV fractions when MLEH and propionic acid were used as carbon sources, and PHBV-MLEH containing 8.9% 3HV was confirmed to have higher thermal stability than standard PHBV (8% 3HV).

Gene Expression of Detoxification Enzymes in Tenebrio molitor after Fungicide Captan Exposure (살진균제인 캡탄 처리 후 갈색거저리의 해독효소 유전자 발현)

  • Jang, Ho am;Baek, Hyoung-Seon;Kim, Bo Bae;Kojour, Maryam Ali Mohammadie;Patnaik, Bharat Bhusan;Jo, Yong Hun;Han, Yeon Soo
    • Korean journal of applied entomology
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    • v.61 no.1
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    • pp.155-163
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    • 2022
  • The application of fungicides is indispensable to global food security, and their use has increased in recent times. Fungicides, directly or indirectly, have impacted insects, leading to genetic and molecular-level changes. Various detoxification mechanisms allow insects to eliminate reactive oxygen species (ROS) toxicity induced by agrochemicals including fungicides. In the present study, we analyzed the mRNA expression levels of detoxifying enzymes in Tenebrio molitor larvae following exposure to non-lethal doses (0.2, 2, and 20 ㎍/µL) of a fungicide captan. Transcripts of peroxidases (POXs), catalases (CATs), superoxide dismutases (SODs), and glutathione-s-transferases (GSTs) were screened from the T. molitor transcriptome database. RT-qPCR analysis showed that TmPOX5 mRNA increased significantly 24 h post-captan exposure. A similar increase was noticed for TmSOD4 mRNA 3 h post-captan exposure. Moreover, the expression of TmCAT2 mRNA increased significantly 24 h post-treatment with 2 ㎍/µL captan. TmGST1 and TmGST3 mRNA expression also increased noticeably after captan exposure. Taken together, these results suggest that TmPOX5 and TmSOD4 mRNA can be used as biomarkers or xenobiotics sensors for captan exposure in T. molitor, while other detoxifying enzymes showed differential expression.

Long-term Clinical Consequences in Patients with Urea Cycle Disorders in Korea: A Single-center Experience (요소회로대사 질환 환자들의 장기적인 임상 경과에 대한 단일 기관 경험)

  • Lee, Jun;Kim, Min-ji;Yoo, Sukdong;Yoon, Ju Young;Kim, Yoo-Mi;Cheon, Chong Kun
    • Journal of The Korean Society of Inherited Metabolic disease
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    • v.21 no.1
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    • pp.15-21
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    • 2021
  • Purpose: Urea cycle disorder (UCD) is an inherited inborn error of metabolism, acting on each step of urea cycle that cause various phenotypes. The purpose of the study was to investigate the long-term clinical consequences in different groups of UCD to characterize it. Methods: Twenty-two patients with UCD genetically confirmed were enrolled at Pusan National University Children's hospital and reviewed clinical features, biochemical and genetic features retrospectively. Results: UCD diagnosed in the present study included ornithine transcarbamylase deficiency (OTCD) (n=10, 45.5%), argininosuccinate synthase 1 deficiency (ASSD) (n=6, 27.3%), carbamoyl-phosphate synthetase 1 deficiency (CPS1D) (n=3, 13.6%), hyperornithinemia-hyperammonemia-homocitrullinuria syndrome (HHHS) (n=2, 9.1%), and arginase-1 deficiency (ARG1D) (n=1, 4.5%). The age at the diagnosis was 32.7±66.2 months old (range 0.1 to 228.0 months). Eight (36.4%) patients with UCD displayed short stature. Neurologic sequelae were observed in eleven (50%) patients with UCD. Molecular analysis identified 37 different mutation types (14 missense, 6 nonsense, 6 deletion, 6 splicing, 3 delins, 1 insertion, and 1 duplication) including 14 novel variants. Progressive growth impairment and poor neurological outcomes were associated with plasma isoleucine and leucine concentrations, respectively. Conclusion: Although combinations of treatments such as nutritional restriction of proteins and use of alternative pathways for discarding excessive nitrogen are extensively employed, the prognosis of UCD remains unsatisfactory. Prospective clinical trials are necessary to evaluate whether supplementation with BCAAs might improve growth or neurological outcomes and decrease metabolic crisis episodes in patients with UCD.