• Title/Summary/Keyword: Transcription Method

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On-off controllable RNA hybrid expression vector for yeast three-hybrid system

  • Bak, Geunu;Hwang, Se-Won;Ko, Ye-Rim;Lee, Jung-Min;Kim, Young-Mi;Kim, Kyung-Hwan;Hong, Soon-Kang;Lee, Young-Hoon
    • BMB Reports
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    • v.43 no.2
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    • pp.110-114
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    • 2010
  • The yeast three-hybrid system (Y3H), a powerful method for identifying RNA-binding proteins, still suffers from many false positives, due mostly to RNA-independent interactions. In this study, we attempted to efficiently identify false positives by introducing a tetracycline operator (tetO) motif into the RPR1 promoter of an RNA hybrid expression vector. We successfully developed a tight tetracycline-regulatable RPR1 promoter variant containing a single tetO motif between the transcription start site and the A-box sequence of the RPR1 promoter. Expression from this tetracycline-regulatable RPR1 promoter in the presence of tetracycline-response transcription activator (tTA) was positively controlled by doxycycline (Dox), a derivative of tetracycline. This on-off control runs opposite to the general knowledge that Dox negatively regulates tTA. This positively controlled RPR1 promoter system can therefore efficiently eliminate RNA-independent false positives commonly observed in the Y3H system by directly monitoring RNA hybrid expression.

Arg243, Invariably Critical for the Transcriptional Activation of Yeast Gcn4p

  • Cho, Gyu-Chull;Lee, Jae-Yung;Kim, Joon
    • Journal of Microbiology
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    • v.37 no.3
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    • pp.154-158
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    • 1999
  • The arginine residue at position 243 (Arg 243) of the yeast transcription factor, Gcn4p, is invariably conserved among bZIP transcription factors. Using site-directed oligonucleotide saturation mutagenesis involving two-step polymerase chain reaction (PCR) amplification, random mutations were successfully introduced at the codon of 243 in the basic domain of Gcn4p. This mutant library was transformed ito Gcn4p defective yeast strain and selected for the transcriptionally active colonies. All colonies which were transcriptionally active had arginines in the codon 243. In this study, the strand preference by Taq polymerase during mutagenesis was also tested. Oligonucleotides were specially designed to test whether or not the polymerase was preferred using the strand as a template. A population of randomly mutated products were cloned into an appropriate vector and characterized by DNA sequencing analysis. Saturation mutagenesis which was performed efficiently by this method revealed a strong bias in terms of strand preference of Taq polymerase by an approximate ratio of 3 to 1 in this study.

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A Study on Optimal Earth-Moon Transfer Orbit Design Using Mixed Impulsive and Continuous Thrust (순간 및 연속 추력을 이용한 지구-달 최적 전이궤도 설계에 관한 연구)

  • No, Tae-Soo;Jeon, Gyeong-Eon
    • Journal of the Korean Society for Aeronautical & Space Sciences
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    • v.38 no.7
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    • pp.684-692
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    • 2010
  • Based on the planar restricted three body problem formulation, optimized trajectories for the Earth-Moon transfer are obtained. Mixed impulsive and continuous thrust are assumed to be used, respectively, during the Earth departure and Earth-Moon transfer/Moon capture phases. The continuous, dynamic trajectory optimization problem is reformulated in the form of discrete optimization problem by using the method of direct transcription and collocation, and then is solved using the nonlinear programming software. Representative results show that the shape of optimized trajectory near the Earth departure and the Moon capture phases is dependent upon the relative weight between the impulsive and the continuous thrust.

Scutellaria baicalensis Georgi(SBG) inhibits Melanin Synthesis in Mouse B16 Melanoma Cells (α-MSH 유도성 멜라닌 합성에 있어서 황금 추출물의 역할과 작용기전 연구)

  • Hong, Sung-Jin;Kim, Kyung-Jun
    • The Journal of Korean Medicine Ophthalmology and Otolaryngology and Dermatology
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    • v.22 no.2
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    • pp.104-117
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    • 2009
  • Objective : Melanin is one of the most important facor in skin color. Melanin protects human skin from ultraviolet radiation otherwise it causes melanin pigmentation. So this experiment is carried out for test whether Scutellaria baicalensis Georgi(SBG) inhibits melanin synthesis and tyrosinase activity in mouse B16 melanoma cells. Method : The melanin synthesis inhibition effects of SBG were examined by in vitro melanin production assay. We assessed inhibitory effects of SBG on melanin contents from B16F1 melanoma cell, on tyrosinase activity(cell and cell free system), effect of SBG on the expression tyrosinase, Microphthalmia-associated Transcription Factor(MITF), Extracellular signal-regulated Kinase(ERK). Result : SBG inhibited melanin synthesis induced $\alpha$-MSH($\alpha$-Melanin Stimulating Hormone) in B16F1. SBG inhibited tyrosinase activity and expression. And SBG down-regulates MITF and stimulated ERK activation in B16F1. Conclusion : According to above results, SBG was improved its suppression effect to the inhibition of melanin synthesis, tyrosinase activation, and tyrosinase promotor activation. So SBG is considered to be used for an strong source of skin whitening effect.

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Imprinted gene Zinc finger protein 127 is a novel regulator of master pluripotency transcription factor, Oct4

  • Kwon, Yoo-Wook;Ahn, Hyo-Suk;Park, Joo-Young;Yang, Han-Mo;Cho, Hyun-Jai;Kim, Hyo-Soo
    • BMB Reports
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    • v.51 no.5
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    • pp.242-248
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    • 2018
  • Induced pluripotent stem cells (iPSCs) show great promise for replacing current stem cell therapies in the field of regenerative medicine. However, the original method for cellular reprogramming, involving four exogenous transcription factors, is characterized by low efficiency. Here, we focused on using epigenetic modifications to enhance the reprogramming efficiency. We hypothesized that there would be a new reprogramming factor involved in DNA demethylation, acting on the promoters of pluripotency-related genes. We screened proteins that bind to the methylated promoter of Oct4 and identified Zinc finger protein 127 (Zfp127), the functions of which have not yet been identified. We found that Zfp127 binds to the Oct4 promoter. Overexpression of Zfp127 in fibroblasts induced demethylation of the Oct4 promoter, thus enhancing Oct4 promoter activity and gene expression. These results demonstrate that Zfp127 is a novel regulator of Oct4, and may become a potent target to improve cellular reprogramming.

Automatic Music Transcription System Using SIDE (SIDE를 이용한 자동 음악 채보 시스템)

  • Hyoung, A-Young;Lee, Joon-Whoan
    • The KIPS Transactions:PartB
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    • v.16B no.2
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    • pp.141-150
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    • 2009
  • This paper proposes a system that can automatically write singing voices to music notes. First, the system uses Stabilized Diffusion Equation(SIDE) to divide the song to a series of syllabic parts based on pitch detection. By the song segmentation, our method can recognize the sound length of each fragment through clustering based on genetic algorithm. Moreover, this study introduces a concept called 'Relative Interval' so as to recognize interval based on pitch of singer. And it also adopted measure extraction algorithm using pause data to implement the higher precision of song transcription. By the experiments using 16 nursery songs, it is shown that the measure recognition rate is 91.5% and DMOS score reaches 3.82. These findings demonstrate effectiveness of system performance.

Nucleic acid-based molecular diagnostic testing of SARS-CoV-2 using self-collected saliva specimens

  • Hwang, Eurim C.;Kim, Jeong Hee
    • International Journal of Oral Biology
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    • v.46 no.1
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    • pp.1-6
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    • 2021
  • Since the outbreak of coronavirus disease 2019 (COVID-2019), the infection has spread worldwide due to the highly contagious nature of severe acute syndrome coronavirus (SARS-CoV-2). To manage SARS-CoV-2, the development of diagnostic assays that can quickly and accurately identify the disease in patients is necessary. Currently, nucleic acid-based testing and serology-based testing are two widely used approaches. Of these, nucleic acid-based testing with quantitative reverse transcription-PCR (RT-qPCR) using nasopharyngeal (NP) and/or oropharyngeal (OP) swabs is considered to be the gold standard. Recently, the use of saliva samples has been considered as an alternative method of sample collection. Compared to the NP and OP swab methods, saliva specimens have several advantages. Saliva specimens are easier to collect. Self-collection of saliva specimens can reduce the risk of infection to healthcare providers and reduce sample collection time and cost. Until recently, the sensitivity and accuracy of the data obtained using saliva specimens for SARS-CoV-2 detection was controversial. However, recent clinical research has found that sensitive and reliable data can be obtained from saliva specimens using RT-qPCR, with approximately 81% to 95% correspondence with the data obtained from NP and OP swabs. These data suggest that self-collected saliva is an alternative option for the diagnosis of COVID-19.

Direct Growth of Patterned-Graphene Using PVP Nanowire Shadow Mask (PVP 나노와이어를 활용한 패턴된 그래핀의 직성장)

  • Eunho Lee;Daesuk Bang
    • Journal of Adhesion and Interface
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    • v.24 no.4
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    • pp.120-123
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    • 2023
  • Graphene, with its exceptional mechanical and electrical properties, has gained significant attention from researchers due to its superior characteristics compared to conventional materials. However, the application of graphene in electronic devices requires a crucial transcription and patterning process, which often introduces numerous defects, substantially impairing its properties. To overcome this limitation and unlock the full potential of graphene for commercial use, there have been various efforts to develop integrated processes for transcription and patterning. In this study, we present a novel growth method that simultaneously achieves precise patterning using polymer nanowires as masks, allowing for the direct growth of graphene. This innovative approach holds promise for realizing advanced electronic components based on nanomaterials in the future.

Detection of Dirofilaria immitis by Reverse Transcription Polymerase Chain Reaction in Canine (역전사중합효소연쇄반응을 이용한 개심장사상충의 검출)

  • 이영준;박진호;권오덕;이주목
    • Journal of Veterinary Clinics
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    • v.16 no.1
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    • pp.177-181
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    • 1999
  • This study was undertaken to clarify the more accurate detecting method of Dirofilaria immitis. Seven dogs, average 7.47 years old, confirmed with Dirofilaria immitis infection by modified Knott's method were used as the experimental animals. cDNA was constructed using oligodT(15) primer after extracting total RNA from the blood of dogs that were confirmed with Dirofilaria immitis infection. As a result of polymerase chain reaction with template using constructed cDNA, the predicted products of a 378 base-pair DNA fragment was amplified. From these results, RT-PCR was more sensitive and effective than modified Knott's method to detect Dirofilaria immitis in dogs.

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