• Title/Summary/Keyword: cell library

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Apoptosis Suppressor에 관련된 유전자 스크린 방법과 동정된 유전자 특성 규명

  • 황규찬;옥도원;권득남;신혜경;김진회
    • Proceedings of the KSAR Conference
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    • 2001.03a
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    • pp.16-16
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    • 2001
  • Apoptosis로 일컬어지는 예정된 세포사멸(programmed cell death)은 개별 세포의 입장에서는 곧바로 사멸을 의미하지만, 정상적인 고등 생물의 입장에서는 개체의 발생과 분화하는데 프로그램된 과정이다. 자발적 세포사멸은 다른 조직에 비해 생식 조직인 난소나 정소에서 복잡한 apoptosis 기작들을 가지리라 사료된다. 본 연구는 Bcl-2 family중 apoptotic protein인 Bax에 대해 suppression하는 유전자를 yeast system을 활용하여 돼지 정소와 난소로부터 각각 cDNA library를 구축한 후 탐색하였다. 탐색에 활용된 cDNA library는 돼지의 정소와 난소로부터 mRNA를 분리하여 yeast vector인 pAD-GAL4-2.1에 구축하였고, 마우스 bax 유전자는 gal 1 promoter의 조절 하에 glucose 배지에서는 유도되지 않고, galactose 배지에서만 선택적으로 Bax를 발현할 수 있는 효모 vector(pL19-bax)를 구축하였다. Bax에 의한 apoptosis suppressor를 탐색하기 위해 우선 효모 W303에 pL19-bax를 transform하여 glucose 배지에서 Bax의 발현을 억제하였다. pL19-bax를 가진 효모에 정소와 난소로부터 구축된 cDNA library를 transform 시키고, transform된 효모는 각각 Bax에 의한 toxicity를 저해하는 유전자를 찾기 위해 스크린되었다. 이러한 방법으로 정소 cDNA library 탐색에서는 5 $\times$ $10^{6}$ transformant중 39개, 난소cDNA library 탐색에서는 2 $\times$ $10^{6}$ transformant중 26개의 콜로니가 생존하였다. 이들 콜로니로부터 유전자를 분리하여 분석해 본 결과 여러 그룹으로 분류할 수 있었다. 각 그룹의 관련 유전자는 protein synthesis/degradation 12종, oxidation/reductation 5종, detoxin/ cell cycle promoter 3종, signal transduction/growth factor 5종, 그리고 알려지지 않은 유전자 9종이었다. 그 중, bax-toxicity inhibition에 강력한 survival phenotype을 가지는 유전자(pSEDL)를 동정하였다. 이것은 T3-4-1 콜로니로부터 분리하였는데 140개 아미노산으로 이루어진 인간 SEDL(GenBank, XM_013096) 유전자와 매우 유사한 homology를 가지며, bax와 관련된 기능은 밝혀져 있지 않다. 이외에도 분리된 유전자에는 NADH, thioreduction, 그리고 cytochrome oxidase와 같은 positive 유전자 군이 크로닝되어, Bax를 이용한 효모에서 apoptosis suppressor에 관련된 유전자를 손쉽게 스크린하는 것이 가능하고, 분리된 유전자의 기능을 예측할 수 있어 지금까지 보고된 유전자 크로닝법 보다는 강력한 수단으로 활용될 수 있다는 사실을 시사하였다. 그러나, ORF에 관계없이 Bax 발현에 저항하는 유전자군이 선발된다든지 하는 문제점은 금후 검토가 필요하리라 사료된다.

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Cloning of cDNA Encoding Putative Cellular Receptor Interacting with E2 protein of Hepatitis C Virus (C형 간염바이러스 E2 단백질에 결합하는 추정 세포수용체 cDNA의 클로닝)

  • 이성락;백재은;석대현;박세광;최인학
    • Journal of Life Science
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    • v.13 no.4
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    • pp.541-550
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    • 2003
  • E2 glycoprotein of hepatitis C virus (HCV) comprises a surface of viral particle together with E1 glycoprotein, and is thought to be involved in the attachment of HCV viral particle to receptor (s) on the permissible cells including hepatocytes, B cells, T cells, and monocytes. We constructed a phage library expressing cellular proteins of hepatocytes on the phage surface, which turned out to be 8.8${\times}$$10^5$ cfu of diversity and carried inserts in 95% of library. We screened both cDNA phage library and 12-mer peptide library to identify the cellular proteins binding to E2 protein. Some intracellular proteins including tensin and membrane band 4.1 which are involved in signal transduction of survival and cytoskeleton organization, were selected from cDNA phage library through several rounds of panning and screening. On the contrary, membrane proteins such as CCR7, CKR-L2, and insulin-like growth factor-1 receptor were identified through screening of peptide library. Phages expressing peptides corresponding to those membrane proteins were bound to E2 protein specifically as determined by neutralization of binding assay. Since it is well known that HCV can infect T cells as well as hepatocytes, we examined to see if E2 protein can bind to CCR7, a member of C-protein coupled receptor family expressed on T cells, using CCR7 transfected tells. Human CCR7 cDNA was cloned into pcDNA3.1(-) vector and transfected into human embryonic kidney cell, 293T, and expressed on the surface of the cell as shown by flow cytometer. Binding assay of E2 protein using CCR7 transfected cells indicated that E2 protein bound to CCR7 by dose-dependent mode, giving rise to the possibility that CCR7 might be a putative cellular receptor for HCV.

A Screen for Dual-protection Molecules from a Natural Product Library against Neuronal Cell Death and Microglial Cell Activation (신경세포 사멸과 미세아교세포활성화 억제 동시 가능 천연물질 탐색 연구)

  • Min, Ju-Sik;Lee, Dong-Seok
    • Journal of Life Science
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    • v.25 no.6
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    • pp.656-662
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    • 2015
  • Natural products and natural product structures play a general and highly significant role in drug discovery and development process because it has various merits and potentials for new drug source that have extensive clinical experience, development time contraction, excellent stability and safety. In several neurological disorders, neuronal death and excessive activation of microglia (neuro-inflammation) are observed. A number of drug discovery-related neuronal cell death and neuro-inflammation was studied from natural products, respectively. However, until now, it has not been possible to study dual-protection molecules recorded in the Natural Product library. In the present study, using the natural product-derived library of the Institute for Korea Traditional Medical Industry, we investigated dual-protective molecules against glutamate (a classical excitatory neurotransmitter)-induced oxidative stress mediated neuronal cell death and LPS-induced excessive activated microglial cells (immune cells of the brain). Chrysophanol, extracted from Rheum palmatum, had dual-protective effects against both glutamate-induced neuronal cell death and LPS-induced NO production, triggering proinflammatory cytokines and microglia activation and resulting in neuroinflammation. Flow-cytometry analysis revealed that chrysophanol had a scavenger effect, scavenging glutamate- and LPS-induced reactive oxygen species (ROS) produced by neuronal and microglial cells, respectively. Based on the present study, chrysophanol may have an important protective role against neuronal cell death and neuroinflammation in the brain. The results may be helpful for studying drug development candidates for treating central nervous system disorders.

Synthesis of 2-Thio-4-aminopyrimidine Derivatives as Anti-cancer Agent

  • Lee, Sang-Hyo;Lee, Jin-Ho
    • Biomedical Science Letters
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    • v.15 no.2
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    • pp.105-112
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    • 2009
  • The screening of the anti-cancer activity of the chemical library provided 2-thio-4-aminopyrimidine as the initial hit. The confirmation of structure and biological effect of hit was performed by synthesis and biological evaluation. The optimization of hit was performed by derivatization of substituents while keeping the core structure. The evaluation of growth inhibitory activity was carried out using SRB assay against 6 human cancer cell lines and human fibroblast. The growth inhibitory activity of compounds showed substituent dependency and more than 5 compounds showed complete growth inhibition of cancer cell lines at 10 ${\mu}M$ concentration. Chemical library screening followed by synthetic modification provided possibility that 2-thio-4-aminopyrimidine can be used as a new scaffold for the development of anti-cancer agent.

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Implementation of Radix-2 structure to reduce chip size (Chip면적 감소를 위한 Radix-2구조 구현)

  • 최영식;한대현
    • Proceedings of the Korean Institute of Information and Commucation Sciences Conference
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    • 1999.05a
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    • pp.407-410
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    • 1999
  • Viterbi decoder is implemented with a Radix-4 architecture at 0.5$\mu\textrm{m}$ process even though the delay time of standard tell is big and it causes a bigger chip size. As process develops, the delay time of standard cells is getting smaller. Therefore, the requirement of speed and chip size is satisfied by using Radix-2 algorithm to implement Viterbi decoder.

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A Study on the Effect of Propagation Delay Time on Critical Time in Storage Elements (기억논리소자에서의 전달지연시간에 의한 Critical Time의 변화 양상 고찰)

  • Joo, Y.J.;Lee, S.H.;Ryoo, J.H.;Lee, S.H.;Sung, Y.K.
    • Proceedings of the KIEE Conference
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    • 1995.07b
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    • pp.922-924
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    • 1995
  • The modeling of accurate timing in storage elements of ASIC cell library was studied. The propagation delay time of clock signal affects the critical time and this can cause malfunction in the chip designed in synchronous. In this paper, an analysis on the effect of input slope of clock signal in timing modeling were carried out. For the first time, in ASIC design, the design guides that can be used in both $0.6{\mu}M$ and $0.8{\mu}m$ design rule were offered, reducing the run time of SPICE and the time of cell library development.

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Analyses of on-the-fly generation of spectral superhomogenization factors for multigroup whole core calculation employing pin-wise slowing-down solutions

  • Seungug Jae;Han Gyu Joo
    • Nuclear Engineering and Technology
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    • v.55 no.3
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    • pp.1084-1096
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    • 2023
  • On-the-fly(OTF) generation of Spectral Superhomogenization(SSPH) factors is analyzed in the multigroup(MG) whole core calculation employing pin-wise continuous energy(CE) slowing-down solutions. The motivation for the work is to avoid the huge computing time required for the generation of a parametrized SSPH factor library(PSSL) which is used to resolve the angular dependency of MG resonance cross sections, and also to exploit the advantage of flexible choice of a MG structure by using CE slowing-down solutions. Two pin-wise CE slowing-down methods, the equivalent Dancoff cell method and the shadowing effect correction method, are evaluated with the OTF SSPH method. The effectiveness of the OTF SSPH method is examined for various simplified and realistic core problems with various MG structures. It is demonstrated that the computing time overhead of this method is negligible whereas the solution accuracy is considerably enhanced.

A Study on the Development of Semi-automated Analog Cell Compiler for MML Library (MML(merged memory logic) 라이브러리 구축을 위한 반자동 아날로그 컴파일러 개발에 관한 연구)

  • 최문석;송병근곽계달
    • Proceedings of the IEEK Conference
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    • 1998.10a
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    • pp.695-698
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    • 1998
  • Today SOC(system on a chip) is a trend in VLSI design society. Especially MML(merged memory Logic) process provides designers with good chances to implement SOC which is consists of DRAM, SRAM, Logic and A/D mixed mode ciruit blocks. Designers need good circuit library which is reliable and easy to tune for specific design. For this need we present semi-automated analog compiler methodology. And we aplied this design methodology to resistor-string DAC design.

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Design of CMOS PLA Using C Language (C언어를 이용한 CMOS PLA의 설계)

  • 차균현;케빈·카플러스
    • Journal of the Korean Institute of Telematics and Electronics
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    • v.21 no.5
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    • pp.61-66
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    • 1984
  • In this paper a custom design of CMOS PLA using procedual language, CHISEL is presented. Library of cells of PLA pieces are formed. A typical PLA is used as a control logic for the protector circuit. NCR's design rules are applied to program CMOS PLA using CHISEL which is a VILI layout language made by extending C language.

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