• Title/Summary/Keyword: the template method

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Site-Specific Mutagenesis on the 32-T and 39-T of E. coli $tRNA^{phe}$ Gene (E. coli $tRNA^{phe}$ 유전자의 32-T와 39-T 염기의 부의 특이적 돌연변이)

  • 김익영;이세영
    • Korean Journal of Microbiology
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    • v.27 no.3
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    • pp.176-180
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    • 1989
  • There are three pseudourdine ($Psi$)bases in the E. coli $tRNA^{phe}$ In order to study the function of the pseudouridine bases in the $tRNA^{phe}$, changes of bases $tRNA^{phe}$ gene to other bases were undertaken by the site-specific mutagenesis. Site-specific mutagenesis of T in the pheW gene, a $tRNA^{phe}$ gene of E. coli, corresponding to the baseat the No.32 position to C and also T corresponding to the base at the No.39 position to C were performed using Kunkel's uracil-containing template method. Identification of mutants were undertaken by the KNA sequencing techniques of the mutated pheW genes and activities of the mutated pheW genes complementing to E. coli NP37 mutant($pheS^{-ts}$) using the recombinant plasmid containing the mutated genes. Neither NP37 harboring pheW gene mutated at No.32 position nor NP37 harboring pheW gene mutated at No.39 position can be grown at non-permissive temperature. The result means that both mutated pheW genes can not complement to E. coli NP37, and that the pseudouridine bases are essential to the activity of the E. coli $tRNA^{phe}$ in vivo.

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Electrochemical Performance of Pitch coated Nano Silicon Sheets / Graphite Composite as Anode Material (피치로 코팅된 Nano Silicon Sheets/Graphite 음극복합소재의 전기화학적 특성)

  • Lee, Tae Heon;Lee, Jong Dae
    • Korean Chemical Engineering Research
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    • v.59 no.4
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    • pp.487-492
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    • 2021
  • In this study, the electrochemical properties of pitch coated silicon sheets/graphite anode materials were investigated. Using NaCl as a template, silicon sheets were prepared through the stöber method and the magnesiothermic reduction methode. In order to synthesize the anode composite, the silicon sheets and graphite were combined with SDBS. The pitch coated silicon sheets/graphite was synthesized using THF as a solvent for the anode material composite. The physical properties of the prepared anode composites were analysed by XRD, SEM, EDS and TGA. The electrochemical performances of the prepared anode composites were performed by the current charge/discharge, rate performance, cyclic voltammetry and EIS tests in the electrolyte LiPF6 dissolved solvents (EC:DMC:EMC=1:1:1 vol%). As the silicon composition of silicon sheets/graphite composite material increased, the discharge capacity also increased, but the cycle stability tended to decrease. The anode material of pitch coated silicon sheets/graphite composite (silicon sheets:graphite=3:7 weight ratio) showed the initial discharge capacity of 1228.8 mAh/g and the capacity retention ratio of 77% after 50 cycles. From these results, it was found that the cycle stability of pitch coated silicon sheets/graphite was improved.

Meal skipping habits and nutritional status among Ghanaian students living in Korea

  • Dzatse, Monica E.;Kim, Eun-kyung;Kim, Hyesook;Kim, Hyejin;Vijayakumar, Aswathy;Chang, Namsoo
    • Journal of Nutrition and Health
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    • v.50 no.4
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    • pp.376-382
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    • 2017
  • Purpose: The consistent rise in the number of foreign students in Korea demands an accurate and detailed investigation into their dietary practices and nutritional status. For these international students, assimilation into new cultures can be stressful. The influence this process may have on dietary behaviors as well as overall health cannot be overlooked. Methods: The researchers in this study sought to investigate the nutritional status and dietary practices of Ghanaian students studying in Korea. A total of 81 Ghanaian male students with an average age of $29.4{\pm}4.0years$ were sampled between May and June 2016. Investigations were carried out on the general characteristics of the participants, their daily food and nutrient intakes using a 24-h dietary recall method, meal skipping practices, and the nutritional quality of their diets based on their meal skipping habits. Results: The study revealed that the daily nutrient intake of subjects did not fully meet the daily recommended nutrient intake (RNI) established by Ghanaian Ministry of Health and the World Health Organization (WHO) and Food and Agriculture Organization (FAO), particularly for energy, calcium, iron, zinc, and vitamin B2 requirements. The Nutrient Adequacy Ratio (NAR) for calcium was very low, posing a deficiency risk to the participants. Meal skipping practices among participants changed significantly after arriving in Korea. The study also compared the diet quality indicators (Nutrient Adequacy Ratio and Mean Adequacy Ratio) of those who skipped meals ${\geq}7times/week$ with those who skipped meals < 7 times/week. Participants who skipped meals ${\geq}7times/week$ were shown to have lower dietary intakes of vitamins B1 and B2 than those who skipped meals < 7 times/week. Conclusion: Ultimately, the study found that meal skipping among Ghanaian students was common, and their daily nutrient intakes did not meet the set dietary standards. Those who skipped meals ${\geq}7times/week$ had lower dietary intakes of vitamins B1 and B2 than those who skipped meals < 7 times/week. Based on our study findings, we recommend that nutrition education should be organized for the Ghanaian student community in Korea using our results as a template to draw up a nutrition program that is culturally appropriate.

Formation of GaN microstructures using metal catalysts on the vertex of GaN pyramids (금속촉매를 이용한 GaN 피라미드 꼭지점 위의 마이크로 GaN 구조 형성)

  • Yun, W.I.;Jo, D.W.;Ok, J.E.;Jeon, H.S.;Lee, G.S.;Jung, S.K.;Bae, S.M.;Ahn, H.S.;Yang, M.
    • Journal of the Korean Crystal Growth and Crystal Technology
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    • v.21 no.3
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    • pp.110-113
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    • 2011
  • In this paper, we propose a new method for the fabrication of GaN microstructures formed only on the vertex of GaN pyramid by using of metal catalysts. GaN pyramidal structures were selectively grown on 3 ${\mu}m$ $SiO_2$ dot patterns followed by thin film deposition of Au and Cr only on the vertex area of the GaN pyramids with precisely controlled photolithography. After the metal deposition, the samples were loaded in the MOVPE reactor for the growth of GaN microstructures for 10 minutes. Temperature for the growth of the GaN microstructures was changed from $650^{\circ}C$ to $750^{\circ}C$. Rod type GaN microstructures were grown in the direction of vertical to the six {1-101} facets and the shape of the GaN microstructures was changed depend on the type of metal.

Optimization of DNA Extraction and PCR Conditions for Fungal Metagenome Analysis of Atmospheric Particulate Matter (대기 입자상물질 시료의 곰팡이 메타게놈 분석을 위한 DNA 추출 및 PCR 조건 최적화)

  • Sookyung Kang;Kyung-Suk Cho
    • Microbiology and Biotechnology Letters
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    • v.51 no.1
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    • pp.99-108
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    • 2023
  • Several challenges arise in DNA extraction and gene amplification for airborne fungal metagenome analysis from a particulate matter (PM) samples. In this study, various conditions were tested to optimize the DNA extraction method from PM samples and polymerase chain reaction (PCR) conditions with primer set and annealing temperature. As a result of comparative evaluation of DNA extraction under various conditions, chemical cell lysis using buffer and proteinase K for 20 minutes and bead beating treatment were followed by using a commercial DNA extraction kit to efficiently extract DNA from the PM filter samples. To optimize the PCR conditions, PCR was performed using 10 primer sets for amplifying the ITS2 gene region. The concentration of the PCR amplicon was relatively high when the annealing temperature was 58℃ with the ITS3tagmix3/ITS4 primer set. Even under these conditions, when the concentration of the PCR product was low, nested PCR was performed using the primary PCR amplicon as the template DNA to amplify the ITS2 gene at a satisfactory concentration. Using the methods optimized in this study, DNA extraction and PCR were performed on 15 filter samples that collected PM2.5 in Seoul, and the ITS2 gene was successfully amplified in all samples. The optimized methods can be used for research on analyzing and interpreting the fungal metagenome of atmospheric PM samples.

Partial Sequencing and Characterization of Porcine DNA Methyltransferase I cDNA

  • Lee, Y.Y.;Kim, M.S.;Park, J.J.;H.Y. Kang;Y.M. Chang;Yoon, J.T.;K.S. Min
    • Proceedings of the Korean Society of Developmental Biology Conference
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    • 2003.10a
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    • pp.84-84
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    • 2003
  • DNA methylation is involved in epigenetic processes such as X-chromosome inactivation, imprinting and silencing of transposons. DNA methylation is a highly plastic and critical component of mammalian development The DNA methyltransferases (Dnmts) are responsible for the generation of genomic methylation patterns, which lead to transcriptional silencing. The maintenance DNA methyltransferase enzyme, Dnmt 1, and the de novo methyltransferase, Dnmt3a and Dnmt3b, are indispensable for development because mice homozygous for the targeted disruption of any of these genes are not viable. The occurrence of DNA methylation is not random, and it can result in gene silencing The mechanisms underlying these processes are poorly understood. It is well established that DNA methylation and histone deacetylation operate along a common mechanistic pathway to repress transcription through the action of methyl-binding domain proteins (MBDs), which are components of, or recruit, histone deacetylase (HDAC) complexes to methylated DNA. As a basis for future studies on the role of the DNA-methyl-transferase in porcine development, we have isolated and characterized a partial cDNA coding for the porcine Dnmt1. Total RNA of testis, lung and ovary was isolated with TRlzol according to the manufacture's specifications. 5 ug of total RNA was reverse transcribed with Super Script II in the presence of porcine Dnmt 1 specific primers. Standard PCRs were performed in a total volume of 50 ul with cDNA as template. Two DNA fragmenets in different position were produced about 700bp, 1500bp and were cloned into pCR II-TOPO according to the manufacture's specification. Assembly of all sequences resulted in a cDNA from 158bp of 5'to 4861bp of 3'compare with the known human maintenance methyltransferase. Now, we are cloning the unknown Dnmt 1 region by 5'-RACE method and expression of Dnmt 1 in tissues from adult porcine animals.

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Development of Raw Material Identification Method of Changnan-jeot and Gaiyang-jeot Using Multiplex PCR and Real-Time PCR (Multiplex PCR과 Real-Time PCR을 이용한 창난젓과 가이양젓 원료 검사법 개발)

  • Choi, Seong Seok;Seo, Yong Bae;Kim, Jong-Oh;Yang, Ji-Young;Shin, Jiyoung;Kim, Gun-Do
    • Journal of Food Hygiene and Safety
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    • v.36 no.4
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    • pp.289-297
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    • 2021
  • In this study, multiplex PCR and real-time PCR were performed on Theragra chalcogramma (walleye pollock), Pangasianodon hypophthalmus (iridescent shark) and their processed foods, such as changnan-jeot and gaiyang-jeot (salted iridescent shark intestine). Species-specific primers for T. chalcogramma and P. hypophthalmus were designed, and genomic DNA was directly extracted from each sample to perform single PCR and multiplex PCR. As a result of PCR, in the case of single PCR, PCR bands of T. chalcogramma (297 bp) and P. hypophthalmus (132 bp) were identified, and in the case of multiplex PCR, it was confirmed that amplification occurred without cross-reaction between T. chalcogramma and P. hypophthalmus. As a result of checking the PCR sensitivity, the concentration of genomic DNA was detected up to 0.1 ng/µL in both single PCR and multiplex PCR. The real-time PCR results showed that the average Ct value of T. chalcogramma was 20.765±0.691, and the average Ct value of P. hypophthalmus sample was 35.719±1.828 in the T. chalcogramma species-specific primers. In the P. hypophthalmus species-specific primers, the average Ct value of the T. chalcogramma sample was 35.996±1.423, and the mean Ct value of the P. hypophthalmus sample was 20.096±0.793. These results demonstrated the significant differences in the efficiency, specificity and cross-reactivity of species-specific primers in real-time PCR. Based on these findings, 7 of changnan-jeot or gaiyang-jeot products were confirmed by multiplex PCR and real-time PCR, and valid results were confirmed in all samples.

Production, Immobilization, and Characterization of Croceibacter atlanticus Lipase Isolated from the Antarctic Ross Sea (남극 로스해에서 분리한 Croceibacter atlanticus균 유래 리파아제의 생산, 고정화, 효소특성 연구)

  • Park, Chae Gyeong;Kim, Hyung Kwoun
    • Microbiology and Biotechnology Letters
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    • v.46 no.3
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    • pp.234-243
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    • 2018
  • The Antarctic Ocean contains numerous microorganisms that produce novel biocatalysts that can have applications in various industries. We screened various psychrophilic bacterial strains isolated from the Ross Sea and found that a Croceibacter atlanticus strain (Stock No. 40-F12) showed high lipolytic activity on a tributyrin plate. We isolated the corresponding lipase gene (lipCA) by shotgun cloning and expressed the LipCA enzyme in Escherichia coli cells. Homology modeling of LipCA was carried out using the Spain Arreo lake metagenome alpha/beta hydrolase as a template. According to the model, LipCA has an ${\alpha}/{\beta}$ hydrolase fold, Gly-X-Ser-X-Glymotif, and lid sequence, indicating that LipCA is a typical lipase enzyme. Active LipCA enzyme was purified fromthe cell-free extract by ammonium sulfate precipitation and gel filtration chromatography. We determined its enzymatic properties including optimum temperature and pH, stability, substrate specificity, and organic solvent stability. LipCA was immobilized by the cross-linked enzyme aggregate (CLEA) method and its enzymatic properties were compared to those of free LipCA. After cross-linking, temperature, pH, and organic solvent stability increased considerably, whereas substrate specificities did not changed. The LipCA CLEA was recovered by centrifugation and showed approximately 40% activity after 4th recovery. This is the first report of the expression, characterization, and immobilization of a C. atlanticus lipase, and this lipase could have potential industrial application.

Real-Time RT-PCR on SAG1 and BAG1 Gene Expression during Stage Conversion in Immunosuppressed Mice Infected with Toxoplasma gondii Tehran Strain

  • Selseleh, Monavar;Modarressi, Mohammad Hossein;Mohebali, Mehdi;Shojaee, Saeedeh;Eshragian, Mohammad Reza;Selseleh, Mina;Azizi, Ebrahim;Keshavarz, Hossein
    • Parasites, Hosts and Diseases
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    • v.50 no.3
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    • pp.199-205
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    • 2012
  • Toxoplasmic encephalitis is caused by reactivation of bradyzoites to rapidly dividing tachyzoites of the apicomplexan parasite Toxoplasma gondii in immunocompromised hosts. Diagnosis of this life-threatening disease is problematic, because it is difficult to discriminate between these 2 stages. Toxoplasma PCR assays using gDNA as a template have been unable to discriminate between an increase or decrease in SAG1 and BAG1 expression between the active tachyzoite stage and the latent bradyzoite stage. In the present study, real-time RT-PCR assay was used to detect the expression of bradyzoite (BAG1)- and tachyzoite-specific genes (SAG1) during bradyzoite/tachyzoite stage conversion in mice infected with T. gondii Tehran strain after dexamethasone sodium phosphate (DXM) administration. The conversion reaction was observed in the lungs and brain tissues of experimental mice, indicated by SAG1 expression at day 6 after DXM administration, and continued until day 14. Bradyzoites were also detected in both organs throughout the study; however, it decreased at day 14 significantly. It is suggested that during the reactivation period, bradyzoites not only escape from the cysts and reinvade neighboring cells as tachyzoites, but also converted to new bradyzoites. In summary, the real-time RT-PCR assay provided a reliable, fast, and quantitative way of detecting T. gondii reactivation in an animal model. Thus, this method may be useful for diagnosing stage conversion in clinical specimens of immunocompromised patients (HIV or transplant patients) for early identification of tachyzoite-bradyzoite stage conversion.

Soccer Game Analysis I : Extraction of Soccer Players' ground traces using Image Mosaic (축구 경기 분석 I : 영상 모자익을 통한 축구 선수의 운동장 궤적 추출)

  • Kim, Tae-One;Hong, Ki-Sang
    • Journal of the Korean Institute of Telematics and Electronics S
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    • v.36S no.1
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    • pp.51-59
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    • 1999
  • In this paper we propose the technique for tracking players and a ball and for obtaining players' ground traces using image mosaic in general soccer sequences. Here, general soccer sequences mean the case that there is no extreme zoom-in or zoom-out of TV camera. Obtaining player's ground traces requires that the following three main problems be solved. There main problems: (1) ground field extraction (2) player and ball tracking and team indentification (3) player positioning. The region of ground field is extracted on the basis of color information. Players are tracked by template matching and Kalman filtering. Occlusion reasoning between overlapped players in done by color histogram back-projection. To find the location of a player, a ground model is constructed and transformation between the input images and the field model is computed using four or more feature points. But, when feature points extracted are insufficient, image-based mosaic technique is applied. By this image-to-model transformation, the traces of players on the ground model can be determined. We tested our method on real TV soccer sequence and the experimental results are given.

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