• Title/Summary/Keyword: PCR product

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Effect of Duck's Feet Derived Collagen Sponge on Skin Regeneration: In Vitro Study (오리발 유래 콜라겐 스펀지의 피부재생 효과: In vitro 연구)

  • Cha, Se Rom;Jeong, Hyun Ki;Kim, Su Young;Kim, Eun Young;Song, Jeong Eun;Park, Chan Hum;Kwon, Soon Yong;Khang, Gilson
    • Polymer(Korea)
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    • v.39 no.3
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    • pp.493-498
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    • 2015
  • For biomaterials for skin regeneration with minimized inflammatory response, high bioactivity and biocompatibility are highly required. Also, it should have a porous microstructure to improve cell adhesion and growth. In this study, we extracted a new collagen source from duck's feet which is by-product, and made the shape of sponges from duck's feet collagen (DC) to compare with DBP and SIS. To analyze physical and chemical property of the scaffold, SEM and FTIR were used. MTT assay was used to measure the attachment and proliferation of NIH/3T3 in the scaffolds. RTPCR was used to evaluate the expression of proinflammatory cytokine. Also, 1,1-diphenyl-2-picrylhydrazyl (DPPH) was used to measure the ability of antioxidant activity. Overall, this study shows that DC scaffold is biocompatible and has good physical property. Additionally, DC scaffold shows the potential as wound healing biomaterials.

Mechanisms of Korean red ginseng and herb extracts(KTNG0345) for anti-wrinkle activity (홍삼 생약 복합물(KTNG0345)의 피부 주름개선에 관한 작용기전)

  • So, Seung-Ho;Lee, Seong-Kye;Hwang, Eui-Il;Koo, Bon-Suk;Han, Gyeong-Ho;Chung, Jin-Ho;Lee, Min-Jung;Kim, Na-Mi
    • Journal of Ginseng Research
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    • v.32 no.1
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    • pp.39-47
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    • 2008
  • UV irradiation causes skin-aging involving coarse wrinkles, thickening, dyspigmentation, and rough skin surface. These phenomena in complex skin tissue is controlled with receptor of cell surface growth factor and cytokine receptors. The activation of receptors induces multiple downstream signaling pathways including expression of MMPs (matrix metalloproteinases). This study was aimed to elucidate the mechanism for anti-wrinkle activity of Korean red ginseng, Torilis fructus and Corni fructus mixture (KTNG0345). In this animal study, we have investigated decreasing effects of Korean red ginseng mixture on MMP-3 synthesis through diminishing $TNF-{\alpha}$ signaling that express MMP-1, -3, and -9. c-Jun and c-fos as a component of transcription factor AP-1 (activator protein-1) were analyzed the expression level using real time PCR and western blotting. c-Jun was decreased dose dependent manner both gene and protein level where as cfos was not changed. In upstream, JNK and PAK was not changed, but p38 was decreased in down stream. MMP-3, final product in this pathway was significantly decreased in dose dependent manner. These results suggest that Korean red ginseng mixture have a anti-wrinkle activity through $TNF-{\alpha}$ mediated MMPs expression pathway.

Screening of Natural Product Libraries for the Extension of Cell Life-span through Immune System (면역시스템을 통한 세포수명연장 천연물질 스크린)

  • Yoo, Bo-Kyung;Kwon, Kisang;Ko, Young Hwa;Kim, Hong Geun;Lee, Seokhyun;Park, Kwan-Ho;Choi, Ji-Young;Kwon, O-Yu
    • Journal of Life Science
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    • v.26 no.3
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    • pp.359-363
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    • 2016
  • We have screened four natural products against 640 single compounds, which shows more two folds gene expression for both endoplasmic reticulum aminopeptidase 1 (ERAP1) and FOXO-family transcription factor (FOXO1). The results were as follows. (±)-Car-3-ene-2,5-dione from Asarum sieboldii Miq. is C10H12O2 molecular formula and the 164 kDa molecular weight. Cinobufagin from Bufonis Venennum is C26H34O6 molecular formula and 442 kDa molecular weight. So far reported main biological function is Na+/K+-ATPase inhibition. Corilagin from Euphorbia pekinensis is C27H22O18 molecular formula and 634 kDa molecular weight. Carbonic anhydrase inhibition is well known its biological function. Corydaline from Corydalis turtschaninovii is C22H27NO4 molecular formula and 369 kDa molecular weight. The main biological function is acetylcholinesterase inhibition. In the short future, four types of natural products will be used in longevity experiments with insects. The results may give one of the clues for studying new drug development candidates of the longevity.

Role of ghrelin in the pancreatic exocrine secretion via mitogen-activated protein kinase signaling in rats

  • Lee, Kyung-Hoon;Lee, Jae-Sung;Wang, Tao;Oh, Jin-Ju;Roh, Sanggun;Lee, Hong-Gu
    • Journal of Animal Science and Technology
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    • v.59 no.7
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    • pp.16.1-16.6
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    • 2017
  • Background: This study was performed to investigate the impact of exogenous ghrelin on the pancreatic ${\alpha}$-amylase outputs and responses of pancreatic proteins to ghrelin that may relate to pancreatic exocrine. Methods: Sprague-Dawley male rats (9 weeks old, $300{\pm}10g$) were injected with ghrelin via intraperitoneal (i.p.) infusion at dosage of 0, 0.1, 1.0 and $10.0{\mu}g/kg$ body weight (BW), respectively. The plasma ghrelin and cholecystokinin (CCK) level were determined using enzyme immunoassay kit; the mRNA expression of ghrelin receptor ($GHSR-1{\alpha}$) and growth hormone (GH) receptor were assessed by reverse transcription PCR; the expressions of pancreatic ${\alpha}$-amylase activity, extracellular-signal-regulated kinases (ERK), phosphorylated extracellular-signal-regulated kinases (pERK) and c-Jun N-terminal kinase (JNK) were evaluated by western blotting; moreover the responses of pancreatic proteins to ghrelin were analyzed using the two-dimensional gel electrophoresis system. Results: The exogenous ghrelin (1.0 and $10.0{\mu}g/kg\;BW$) elevated the level of plasma ghrelin (p < 0.05), and suppressed the expression of pancreatic ${\alpha}$-amylase at a dose of $10.0{\mu}g/kg\;BW$ (p < 0.05). No difference in the level of plasma CCK was observed, even though rats were exposed to any dose of exogenous ghrelin. In addition, a combination of western blot and proteomic analysis revealed exogenous ghrelin ($10.0{\mu}g/kg\;BW$) induced increasing the JNK and ERK expressions (p < 0.05) and four proteins such as Destrin, Anionic trypsin-1, Trypsinogen, and especially eukaryotic translation initiation factor 3 in rat pancreas. Conclusions: Taken together, exogenous ghrelin by i.p. infusion plays a role in the pancreatic exocrine secretion via mitogen-activated protein kinase signaling pathway.

Cytological analysis of pregnancy-associated plasma protein-A expression in porcine neonatal testis (미성숙 돼지 정소에서 pregnancy-associated plasma protein-A의 발현의 세포학적 분석)

  • Kim, Ji-youn;Oh, Keon Bong;Byun, Sung June;Ock, Sun-A;Lee, Hwi-Cheul;Hwang, Seong-Su;Park, SangHyun;Ha, Wootae;Woo, Jae-Seok;Song, Hyuk
    • Journal of Embryo Transfer
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    • v.33 no.3
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    • pp.177-183
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    • 2018
  • The identification of biomarkers of a living tissues is essentially required to understand specific functions of the cells. In previous study, we reported IGFBP 3 as one of the putative biomarkers, by showing specific expression at porcine spermatogonial stem cells (SSCs) of early stage of porcine testis. In this study, we analyzed the expression of seven members of IGFBP family (IGFBPs) in SSCs and histological expression pattern of pregnancy-associated plasma protein-A (PAPP-A), which plays a role on the growth promoting enzyme by cleavage of IGFBPs in testis of 5 days old pig. RT-PCR analysis showed that IGFBP 1, 2, 3, 4, and 6 were expressed at high level specifically in porcine SSCs compared with whole testis. We performed immunohisotochemical staining of testis sections with PAPP-A and protein gene product 9.5 (PGP9.5) which are the known biomarkers for SSCs. We were not able to find co-expression of PAPP-A and PGP9.5; PAPP-A was expressed only in Sertoli cells and PGP9.5 expression was confirmed in spermatogonium. Additionally, we were able to confirm the GATA4 expression in Sertoli and Leydig cells as a regulator of Sertoli cell function was not detected PGP9.5 expressing cells, indicating indirect evidence of that cytolocalization of PAPP-A expression is limited in Sertoli cells. These results suggested that the PAPP-A expressed in Sertoli cells may play role on regulation of development and differentiation of testicular cells through the IGF axis in neonatal porcine testis.

Monitoring of Raw Materials for Commercial Home Meal Replacement Products Using DNA Barcode Information (DNA 바코드를 이용한 가정간편식 제품의 원재료 모니터링 연구)

  • Yu, Yeon-Cheol;Hong, Yewon;Kim, Jung Ju;Lee, Dong Ho;Kim, Hyung Soo;Moon, Guiim;Park, Eun Mi
    • Journal of Food Hygiene and Safety
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    • v.35 no.3
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    • pp.234-242
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    • 2020
  • In this study, we monitored the raw materials in home-meal replacement (HMR) products, which have shown more than 63% growth in market size for two years. A total of 89 HMR products were purchased and the DNA barcodes of 112 raw materials in the product samples were analyzed. In order to identify the raw material species, a primer set specific for the 16S ribosomal RNA region of each raw material species was amplified. The amplicon was purified and sequenced, and then used to perform a BLAST search provided by the National Institutes of Health (NIH). The species of the raw material was determined by comparing the nucleotide sequences of the species registered in GenBank with identity and match score. Twenty-four species and three genera were identified from 112 raw materials. Three genera were identified at the genus level because a large number of species belonging to the same genus exist within 98% of the identity criteria. The results of the determination were compared with the available raw materials suggested in the Korea Food Code to determine the Korean name and availability of the foods. Six non-listed species were determined to be edible according to information provided by influential domestic and foreign organizations.

Cloning and Characterization of the Lactococcus lactis subsp. lactis ATCC 7962 pts HI Operon

  • Kim, Tea-Youn;Park, Rae-Jun;Chang, Hae-Choon;Chung, Dae-Kyun;Lee, Jong-Hoon;Lee, Hyong-Joo;Kim, Jeong-Hwan
    • Journal of Microbiology and Biotechnology
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    • v.10 no.6
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    • pp.829-835
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    • 2000
  • The ptsH and ptsI genes of Lactococus lactis subsp. lactis ATCC 7962 (L. lactis 7962), encoding the general proteins of phosphotransferase system (PTS) components, HPr and enzyme I, respectively, were cloned and characterized. A 1.3 kb PCR product was obtained using a primer set that was hybridized to the internal region of the L. lactis 7962 pts HI genes and then subcloned into a low-copy number vector, pACYC184. The 5' upstream and 3' downstream region from the 1.3 kb fragment were subsequently clone using the chromosome walking method. The complete ptsHI operon was constructed and the nucleotide sequences determined. Two ORFs corresponding to HPr (88 amino acids) and enzyme I (575 amino acids) were located. The ptsHI genes of L. lactis 7962 showed a very high homology (84-90%) with those genes from other Gram-positive bacteria. A primer extension analysis showed that the transcription started at either one of two adjacent bases upstream of the start codon. Using a Northern analysis, two transcripts were detected; the first, a 0.3 kb transcript corresponding to ptsH and the second, a 2 kb transcript corresponding to ptsH and ptsI. The transcription level of ptsH was higher than that of ptsI. The concentration of the ptsH transcript in cells grown on glucose was similar to that in cells grown on lactose, yet higher than that in cells grown on galactose. The ptsI transcript was scarcely detected in cell grown on lactose or galactose. The ptsI transcript was scarcely detected in cells grown on lactose or galactose. The results of a sequence analysis and Northern blot confirmed that the ptsH and ptsI genes of L. lactis 7962 were arranged in an operon like other known ptsHI genes and the expression of the ptsHI genes was regulated at the transcriptional level in response to the carbon source.

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The Use of Fibroin Light Chain Gene Sequence for the Genetic Marker of the Silkworm Races

  • Park, Kwang-Ho;Kang, Seok-Woo;Kang, Pil-Don;Goo, Tea-Won;Hwang, Jae-Sam;Yun, Eun-Young;Lee, Sang-Mong;Sohn, Hung-Dae;Jin, Byung-Rae
    • International Journal of Industrial Entomology and Biomaterials
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    • v.6 no.1
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    • pp.45-48
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    • 2003
  • We have previously cloned and characterized the complete fibroin L-chain gene from one of the silkworm races Baekok-Jam (Bombyx mori) and found two variable regions (I, intron 2 ~ exon 3; II, intron 6) with the primer sets designed to cover these variable regions. We tested the utility of these regions as genetic markers among silkworm races. For the purpose of study, Japanese race (Jam 123), Chinese race (Jam 124) and their F$_1$hybrid Baekok-Jam were used. The PCR product size of region I was 787 bp in Jam 123, 770 bp in Jam 124 and 768 bp in Baekok-Jam. The size of region II was 470 bp in Jam 123, 428 bp in Jam 124 and 429 Up in Baekok-Jam. In the extended experiment, Jam 125 (Japanese race), Jam 126 (Chinese race) and their F$_1$hybrid Daeseong-Jam were also analyzed. The sizes of region I and II in Jam 125, Jam 126 and Daeseong-Jam were similar to those of Jam 123, Jam 124 and Baekok-Jam. DNA sequence divergence between the two geographic races of Jam 123 or Jam 125 and Jam 124 or Jam 126 was substantial. The result suggests that the fibroin L-chain gene of F$_1$hybrids were inherited from Chinese races. These results are concordant with cocoon shapes of tested animals, and suggested that Baekok-Jam or Daeseong-jam is more closely related to Jam 124 or Jam 126 than to Jam 123 or Jam 125. Taken these data together, the primer sets designed from two variable regions of fibroin L-chain gene would be highly useful, as the genetic markers for silkworm races, at least in Japanese and Chinese races, although an extended study including more geographic races is required.

Studies on the safety of Brucella abortus RB51 vaccine I. Comparison of the biochemical and genetic characteristics of Brucella abortus RB51 vaccine strains (부루세라백신(RB51)의 안전성에 관한 연구 I. Brucella abortus RB51 백신균주의 생화학적 및 유전학적 성상비교)

  • Kim, Jong-man;Woo, Sung-ryong;Lee, Ji-youn;Jung, Suk-chan;Kang, Seung-won;Kim, Jong-yeom;Yoon, Yong-dhuk;Cho, Sang-nae;Yoo, Han-sang;Olsen, Steven C.
    • Korean Journal of Veterinary Research
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    • v.40 no.3
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    • pp.533-541
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    • 2000
  • Biochemical and genetic analysis were carried out to investigate the potential recovery of pathogenecity or related mutations of Brucella abortus RB51 vaccine strains. RB51 strains were recovered from commercial vaccines, including related seed stocks from private companies in Republic of Korea, strain from USA, a reference strain from C university and a field isolate (Daehungjin) from aborted dairy cow after RB51 vaccination were compared with two identified virulent wild strains (S2308 and a field strain isolated from dairy cow in Korea) at the same conditions. All the strains examined, except identified pathogenic strains, revealed the identical characteristics to the original RB51 in biochemical properties, antigen and bacteriophage typing. Outer membrane protein (OMP) profiles from strains of RB51 showed the same patterns with standard RB51 in SDS-PAGE. In addition, Western blotting with the brucella specific monoclonal antibody also indicated that all the vaccine strains were completely deficient in their LPS compared to the pathogenic Br abortus strains. The differences in DNA structures among strains were also possible to detect after PCR. All vaccine strains, except S19, S1119-3, S1075, S544 and Br suis, were amplified a 178bp DNA fragment of eri-gene, and 364bp of IS711 elements. In contrast, 498bp DNA product was only found with Br abortus. Overall evidences in the present study confirmed that the RB51 strains for vaccine production in Korea did not originated from the phenomena of possible recovery of pathogenicity or related to any potential mutation event at all.

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Structure Analysis of pmcABCDEFT Gene Cluster for Degradation of Protocatechuate from Comamonas sp. Strain DJ-12 (Comamonas sp. Strain DJ-12로부터 Protocatechuate의 분해에 관여하는 pmcABCDEFT 유전자군의 구조 분석)

  • Kang Cheol-Hee;Lee Sang-Mhan;Lee Kyoung;Lee Dong-Hun;Kim Chi-Kyung
    • Korean Journal of Microbiology
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    • v.41 no.3
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    • pp.195-200
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    • 2005
  • Comamonas sp. strain DJ-12 is a bacterial isolate capable of degrading of 4-chlorobiphenyl (4CB) as a carbon and energy source. The degradation pathway was characterized as being conducted by consecutive reactions of the meta-degradation of 4CB, hydrolytic dechlorination of 4-chlorobenzoate (4CBA), hydroxylation of 4-hydroxybenzoate, and meta-degradation of protocatechuate to product TCA metabolites. The 6.8 kb fragment from the chromosomal DNA of Comamonas sp. strain DJ-12 included the genes encoding for the meta-degradation of PCA; the genes of protocatechuate 4,5-dioxygenase alpha and beta subunits (pmcA and pmcB), 4-carboxy-2-hydroxymuconate-6-semialdehyde dehydrogenase (pmcC), 2-pyrone-4,6-dicarboxylate hydrolase (pmcD), 4-oxalomesaconate (OMA) hydratase(pmcE), 4-oxalocitramalate (OCM) aldolase (pmcF), and transporter gene (pmcT). They were organized in the order of pmcT-pmcE-pmcF-pmcD-pmcA-pmcB-pmcC. The amino acid sequences deduced from the nucleotide sequences of pmcABCDEFT genes from Comamonas sp. strain DJ-12 exhibited 94 to $98\%$ homologies with those of Comamonas testosteroni BR6020 and Pseudomonas ochraceae NGJ1, but only 52 to $74\%$ with homologies Sphingomonas paucimobilis SYK-6, Sphingomonas sp. LB126, and Arthrobacter keyseri 12B.