• Title/Summary/Keyword: 중합 효소 연쇄 반응

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GENE EXPRESSION ANALYSIS OF THE DENTAL PULP IN HEALTHY AND CARIES TEETH (치아 우식증에 따른 치수내 유전자 발현 변화에 관한 분석)

  • Oh, So-Hee;Kim, Jong-Soo
    • Journal of the korean academy of Pediatric Dentistry
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    • v.37 no.3
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    • pp.275-287
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    • 2010
  • Deep caries may induce pulpitis and the pulpal tissue interacts with microbial invasion. The immune response to protect the pulpal tissue can be mediated by cellular signal molecules produced by the pulpal cells. The understanding of these processes is important to find future therapeutic method for the diseased pulp. The pulp tissue from sound teeth was set as control group (n=30) and the pulp tissue from decayed teeth was set as test group (n=30). Total RNA was extracted from the pulp of each group and it was used for cDNA microarray and reverse transcriptase-polymerase chain reaction(RT-PCR). The expression of TGF-${\beta}1$ was studied by immunohistochemistry. The results were as follows: 1. cDNA microarray analysis identified 520 genes with 6-fold or greater difference in expression level with 143 genes more abundant in health and 377 genes more abundant in disease. 2. The RT-PCR analysis was done for randomly selected 14 genes and the results supported the result of cDNA microarray assay. 3. TGF-${\beta}1$ was highly expressed in the carious pulp and it was found in odontoblast by immunohistochemistry. In conclusion, many cytokines were found to be significantly changed their expression in the diseased pulp(/M/>1.6).

The Effect of ${\Delta}^{12}PGJ2$ and $PPAR{\Gamma}$ Agonist on the Proliferation and Differentiation of Osteoblast ((${\Delta}^{12}PGJ_2$ 및 PPAR 감마 길항체가 조골세포의 증식 및 분화에 미치는 효과)

  • Heo, Jeong-Mi;Kim, Kyoung-Wha;Chung, Kyoung-Wook;Lee, Hye-Joon;Rhyu, In-Chul;Ku, Young;Chung, Chong-Pyoung
    • Journal of Periodontal and Implant Science
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    • v.35 no.1
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    • pp.133-152
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    • 2005
  • 1. 목적 Prostaglandin은 치주질환과 관련된 국소적 골 대사에 중요한 역할을 한다. ${\Delta}^{12}PGJ_2$는 생체 내에서 혈장의 존재 하에 형성되는 천연 $PGD_2$ 대사산물이며 peroxisome- proliferator에 의해 활성화되는 감마 수용체 (PPAR ${\Gamma}$)에 대해 높은 친화성을 갖는 리간드로서 핵 수용체군에 속하는 전사조절인자이다. 이 연구의 목적은 골화 과정에서 ${\Delta}^{12}PGJ_2$의 역할을 규명하기 위해, 조골세포주의 증식과 분화에 미치는 영향과 그에 관련된 세포기전을 조사하는 데에 있다. 2. 방법 인간 골육종세포주인 Saos-2 (ATCC.HTB 85)와 쥐의 조골세포주 (MC3T3-E1)를 배양한 후 실험군에 농도가 각각 $10^{-5}$, $10^{-6}$, $10^{-7}$, $10^{-8}$, $10^{-9}$ 몰인 ${\Delta}^{12}PGJ_2$와 ciglitazone (합성 PPAR 감마 길항체)를 첨가하였다. 조골세포에서 PPAR 감마의 발현을 관찰하기 위해 역전사효소-중합효소연쇄반응(RT-PCR)을 특정한 primer를 이용하여 시행하였다. 세포 증식은 1일, 2일, 3 일째에 MIT 분석법으로 측정하였고, 2 일째에 알칼리성 인산효소 (ALPase) 생산을 측정하였다. 위의 결과에서 얻은 적정한 농도에서 다양한 조골세포 분화의 표지자들-제 1 형 교원질, 알칼리성 인산효소, osteopontin 및 bone sialoprotein-에 대한 간이 정량적 역전사효소-중합효소연쇄반응 (semiquantitative RT-PCR)을 실시하였으며 골결절 형성에 대한 효과를 알아보고자 석회화 분석도 시행하였다. 3. 결과 ${\Delta}^{12}PGJ_2$와 ciglitazone 모두 Saos-2 세포주의 증식을 촉진시켰다 .$10^{-8}$ 몰의 ${\Delta}^{12}PGJ_2$$10^{-6}$몰의 ciglitazone을 첨가한 실험군을 대조군과 비교했을 때, 시간에 비례하여 세포 증식률이 증가되었다. 알칼리성 인산효소의 활성화 검사에서도 증식률에서와 유사한 결과를 보여주었다. 간이 정량적 RT-PCR에서는 ${\Delta}^{12}PGJ_2$로 처리한 군의 경우 제 1 형 교원질, 알칼리성 인산효소, osteopontin, 그리고 bone sialoprotein의 상대적 mRNA 수준이 유의하게 높았다. 석회화 분석에서는 MC3T3-E1 세포를 $10^{-6}$ 몰의 ${\Delta}^{12}PGJ_2$로 처리한 군과 $10^{-5}$ 몰의 ciglitazone으로 처리한 군에서 현저한 골결절 형성을 보였다. 이러한 결과들은 ${\Delta}^{12}PGJ_2$가 유용한 골 유도물질이 될 수 있으며 또한 그 작용기전이 PPAR 감마-의존형 경로와 연관되어 있음을 보여준다.

PDMS/Glass Serpentine Microchannel Chip for PCR with Bubble Suppression in Sample Injection (시료주입시 기포발생이 억제된 반응조 형태의 중합효소연쇄반응용 PDMS/유리 바이오칩)

  • Cho Chul-Ho;Cho Woong;Hwang Seung-Yong;Ahn Yoo-Min
    • Transactions of the Korean Society of Mechanical Engineers A
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    • v.30 no.10 s.253
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    • pp.1261-1268
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    • 2006
  • This paper reports low-cost microreactor $(10{\mu}{\ell})$ biochip for the DNA PCR (polymerase chain reaction). The microbiochip $(20mm{\times}28mm)$ is a hybrid type which is composed of PDMS (polydimethylsiloxane) layer with serpentine micochannel $(360{\mu}m{\times}100{\mu}m)$ chamber and glass substrate integrated with microheater and thermal microsensor. Undesirable bubble is usually created during sample loading to PMDS-based microchip because of hydrophobic chip surface. Created bubbles interrupt stable biochemical reaction. We designed improved microreactor chamber using microfluidic simulation. The designed reactor has a coner-rounded serpentine channel architecture, which enables stable injection into hydrophobic surface using micropipette only. Reactor temperature needed to PCR reaction is controlled within ${\pm}0.5^{\circ}C$ by PID controller of LabVIEW software. It is experimentally confirmed that SRY gene PCR by the fabricated microreactor chip is performed for less than 54 min.

Development of Detachable PDMS/Glass PCR-Chip and It's Application to Detection of Male Infertility (분리식 PDMS/유리 중합효소연쇄반응칩 개발 및 유전적 남성불임 검사에의 응용)

  • Ju, Jin-Kyoung;Hwang, Seung-Young;Ahn, Yoo-Min
    • Transactions of the Korean Society of Mechanical Engineers A
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    • v.32 no.4
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    • pp.371-377
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    • 2008
  • Our precedent study has reported glass-PDMS (polydimethylsiloxane) based biochip for the gene PCR (polymerase chain reaction). To prevent the contamination of bio sample, the once used biochip must not be used repeatedly. However, the fabrication cost of microheater and microsensor of the biochip was not cheap to use it as a disposable chip. This paper proposes new PCR-chip where the glass substrate integrated with the microheater and microsensor is detachable from the reaction chamber where the sample is injected. That makes it possible to reuse the glass substrate repeatedly. The performance of the proposed detachable PCR-chip was compared with that of the precedent monolithic PCR-chip. The results showed that the SRY (sex determining Y chromosome) gene PCR was successfully performed in the detachable chip compared with the monolithic chip. However, the more efforts to improve the efficiency of surface treatment of PDMS chip are needed to increase the possibility of applying the detachable chip to the detecting of male infertility.

A Study on the Validation system of Detection for Biological Agents Using Real-Time PCR (실시간 중합효소 연쇄반응을 활용한 생물작용제 검증시스템 연구)

  • Cha, Younggil;Koo, Bonwoo;Kim, Seongjoo;Kim, Namil;Park, Hanoh
    • Journal of the Korea Institute of Military Science and Technology
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    • v.20 no.5
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    • pp.726-732
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    • 2017
  • Bacillus anthracis, Vibrio cholerae, Variola virus and Shigella dysenteriae are classified as category A and B biological weapons. In this study suggest that 4 genes of Bacillus anthracis, 2 genes of Vibrio cholerae, 1 gene of Variola virus and 1 gene of Shigella dysenteriae were detective 50~500 fg of target DNA per reaction using real-time PCR based assay. Also analytical specificity did not show any cross-reactivity with other related bacteria. Reliable and one reaction could be effective early diagnostic and treatment for detection of unknown samples.

Isolation of Myxobacteria Carrying Soraphen Biosynthetic Gene Clusters (Soraphen 생합성 유전자군을 갖는 점액세균의 분리)

  • Lee, Cha-Yul;Hyun, Hye-Sook;Cho, Kung-Yun
    • Microbiology and Biotechnology Letters
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    • v.37 no.1
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    • pp.10-16
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    • 2009
  • Polymerase chain reactin (PCR) could be a simple way to screen new microbial strains producing useful secondary metabolites if their biosynthetic genes are known and candidate strains to be screened are available. In this study, we have screened two myxobacterial strains, KYC3047 and KYC3076, carrying genes appeared to be biosynthetic genes of soraphen A, a potent antifungal substance, out of 50 cellulose degrading myxobacteria using PCR. The two strains were identified as Sorangium cellulosum based on morphological, physiological, and molecular biological characteristics. Both of the strains produced substances having strong antifungal activities as expected against Candida albicans, a causative agent of candidiasis, and Colletotrichum acutatum, a causative agent of anthracnose on pepper.

Allele Frequency of the Short Tandem Repeat Locus Human Lipoprotein Lipase(LPL) Gene by Polymerase Chain Reaction in the Korean Population (한국인에서 중합효소연쇄 반응법에 의한 STR 유전좌위 LPL의 유전자빈도 검색)

  • Na, Yun-Ju;Hur, Woong;Yoon, Chang-Lyuk
    • Journal of Oral Medicine and Pain
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    • v.22 no.2
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    • pp.253-260
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    • 1997
  • 한국인 집단에서 개인식별의 기초자료로 활용하고자 한국인 201명을 대상으로 STR 유전좌위 중 하나인 LPL 유전좌위의 유전자 빈도 및 유전자형 분포를 구하였다. 혈액으로부터 추출한 핵 DNA를 중합효소연쇄반응으로 증폭시키고 폴리아크릴아마이드 겔 상에서 전기영동하여 은염색한 후 관찰하여 다음의 결과를 얻었다. 1. 한국인 집단 201명의 LPL 유전자에서 5개의 대립유전자, 7개의 유전자형을 검출하였으며, 이형접합도는 50.7%로 나타났고 대립 유전자다양성 (allelic diversity value)은 0.454, 개 인식 별력 (PD)은 0.674를 보였다. 2. 대립 유전자 및 유전자빈도는 9, 10, 11, 12, 13 대립 유전자에서 각각 0.020, 0.714, 0.100, 0.164, 0.002로 나타났으며, 대립유전자 7, 8, 14는 관찰되지 않았다. 이상의 결과를 볼 때 한국인 집단에서 STR LPL유전좌위의 유전자빈도는 친자감정 등 개인식별에 유용하게 사용할 수 있으나 감정실무에 응용시 다수의 STR유전좌위 및 VNTR유전좌위의 분석을 병행하여야 할 것으로 사료된다.

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p53 Mutations in Ewing's Sarcoma (유잉육종의 p53 돌연변이)

  • Bae, Dae-Kyung;Sun, Seung-Deok
    • The Journal of the Korean bone and joint tumor society
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    • v.6 no.4
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    • pp.143-151
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    • 2000
  • Purpose : The p53 tumor suppressor gene is one of the most frequently altered genes in human malignancies. We try to explore the implication of p53 alteration in Ewing's sarcoma. Materials and Methods : We analyzed 35 paraffin blocks to explore the deletion and sequence alterations of p53. Results : Quantitative PCR analysis showed that 2 tumors showed a homozygous deletion of the gene. Mutational analysis of exons 4 to 9 of p53 by PCR-SSCP revealed that 3 tumors carry sequence alterations in exons 5 or 8, and DNA sequencing analysis identified missense point mutations. Conclusion : Taken together, our data demonstrate that p53 is genetically altered in a small fraction of Ewing's sarcoma.

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