• 제목/요약/키워드: Agarose gel electrophoresis

검색결과 260건 처리시간 0.031초

다양한 정제방법에 의한 전기영동용 한천유래 아가로즈의 제조 및 DNA분리 특성 (Preparation of Agarose from Gelidium amansii for Gel Electrophoresis using Various Purification Methods and Its Resolution Characteristics for DNA)

  • 도정룡;오세욱
    • 한국식품과학회지
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    • 제31권1호
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    • pp.110-114
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    • 1999
  • 제주도산 우뭇가사리로부터 agar를 제조하여 분석용 등급의 agarose 제조를 위하여 cetylpyridium chloride (CPC), polyethylene glycol (PEG), chitosan을 처리하여 agaropectin을 제거하여 agarose를 분리, 정제하였다. 3가지 방법중 chitosan을 처리하여 정제한 agarose는 황산기 함량이 가장 낮았으며, electroendosomosis도 가장 낮은 수치를 나타내어 분석용 agarose 제조를 위한 가장 효율적인 방법으로 나타났으며 실제 DNA 전기영동 실험에서도 가장 우수한 resolution을 나타내었으며 교차면역 전기영동에서도 시판되고 있는 제품에 비하여 품질이 양호함을 알 수 있었다.

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吸着 크로마토그라피法에 의한 사람 ALpha-Fetoprotein의 分離 (Affinity Purification of Human Alpha-Fetoprotein)

  • Kang, Shin-Sung;Bang, Ok-Sun;Park, Tae-Kyu
    • 한국동물학회지
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    • 제29권4호
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    • pp.283-293
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    • 1986
  • 사람의 alpha-fetoprotein(AFP)에 대한 모노클론 항체의 생산 및 분석을 위하여, 태아조직을 재료로 추출법, DEAE-cellulose 및 concanavalin A-Sepharose, Cibacron blue F3GA-agarose, immunoadsorbent column등의 흡착크로마토그라피법에 의해 AFP를 분리하였다. 총 534g의 태아조직에서 AFP의 量은 8.76 mg으로서 순수분리되었음을 확인하였다.

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제주산 우뭇가사리(Gelidium amansii)로부터 DMSO 추출과 EDTA 수세법에 의한 상용화 아가로스 제조 (Preparation of Commercial Agarose from Jeju Seaweed, Gelidium amansii using DMSO Extraction and EDTA Washing)

  • 강태환;이승홍;백종석;강병식;이정석;이남호;전유진
    • 한국수산과학회지
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    • 제44권6호
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    • pp.635-643
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    • 2011
  • Agar was prepared from Gelidium amansii collected from Jeju Island, South Korea. This agar preparation has high gel strength and low sulfate content compared with G. amansii agar from Morocco. Accordingly, agarose was made from the Jeju agar through the consecutive refining processes of dimethyl sulfoxide (DMSO) extraction and ethylene diamine tetra acetic acid (EDTA) washing. The physicochemical properties of the resulting agarose were compared with those from agarose prepared using only DMSO extraction. Consecutive DMSO extraction and EDTA washing more strongly affected the physicochemical properties of the agarose (purified agarose) compared with the use of DMSO extraction alone. These properties were similar to those of commercial agarose used for electrophoresis. In DNA electrophoresis, the separation and movement speed of the purified agarose were similar to those of the commercial agarose. In a $^{13}C$ NMR analysis, the purified agarose exhibited the same carbon peak as the commercial agarose. When observed under scanning electron microscopy, the agar had an even and smooth surface without irregularities or pores, and the purified agarose had a wide surface area with a large number of pores; the commercial agarose had an irregular surface that would allow the solvent to easily permeate. These results illustrate that the physicochemical properties of agarose prepared from DMSO extraction and EDTA washing were more effective than those observed after DMSO extraction alone; thus, these processes used in succession will be useful in agarose industries.

DNA damage와 Apoptosis를 정량화하는 단세포전기영동법 (Single Cell Gel Electrophoresis (comet assay) to Detect DNA Damage and Apoptosis in Cell Level)

  • 류재천;김현주;서영록;김경란
    • 한국환경성돌연변이발암원학회지
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    • 제17권2호
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    • pp.71-77
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    • 1997
  • The single cell gel electrophoressis(SCGE) assay, also known as the comet assay, is a rapid, simple, visual and sensitive technique for measuring and analysing DNA breakage in mammalian cells. The SCGE or comet assay is a promising test for the detection of DNA damage and repair in individnal cells. It has widespread potential applications in DNA damage and repair studies, genotoxicity testing and biomonitoring. In this microgel electrophoresis technique, cells are embedded in agarose gel on microscope slides, iysed and electrophoresed under alkaline conditions. Cells with increased DNA damage display increased migration of DNA from the nucleus towards the anode. The length of DNA migration indicates the amount of DNA breakage in the cell. The comet assay is also capable of identifying apoptotic cells which contain highly fragmented DNA. Here we review the development of the SCGE assay, existing protocols for the detection and analysis of comets, the relevant underlying principles determining the behaviour of DNA and the potential applications of the technique.

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A Method for Direct Application of Human Plasmin on a Dithiothreitol-containing Agarose Stacking Gel System

  • Choi, Nack-Shick;Chung, Dong-Min;Yoon, Kab-Seog;Maeng, Pil-Jae;Kim, Seung-Ho
    • BMB Reports
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    • 제38권6호
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    • pp.763-765
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    • 2005
  • A new simplified procedure for identifying human plasmin was developed using a DTT copolymerized agarose stacking gel (ASG) system. Agarose (1%) was used for the stacking gel because DTT inhibits the polymerization of acrylamide. Human plasmin showed the lowest activity at pH 9.0. There was a similar catalytically active pattern observed under acidic conditions (pH 3.0) to that observed under alkaline conditions (pH 10.0 or 11.0). Using the ASG system, the primary structure of the heavy chain could be established at pH 3.0. This protein was found to consist of three fragments, 45 kDa, 23 kDa, and 13 kDa. These results showed that the heavy chain has a similar structure to the autolysed plasmin (Wu et al., 1987b) but there is a different start amino acid sequence of the N-termini.

BSH 존재시 중성자 및 ${\gamma}$-ray 조사에 따른 plasmid DNA의 손상 (Plasmid DNA damage by neutron and ${\gamma}$-ray in the presence of BSH)

  • 천기정;서원숙
    • Journal of Radiation Protection and Research
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    • 제31권2호
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    • pp.65-68
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    • 2006
  • 본 연구에서는 boron 화합물인 BSH(Boron Sulfhydryl Hydride)의 농도와 중성자 및 감마선 방사선 조사선량에 따른 plasmid DNA의 손상 정도를 관찰하였다. Plasmid는 pBR 322(2870 bp)와 ${\Phi}X174$ RF DNA(5386 bp)를 사용하였고 조사 후 DNA의 손상 정도는 agarose gel 전기영동 상에서 관찰하였다. 중성자 조사에서는 plasmid DNA의 손상 정도는 BSH의 농도 및 조사 선량이 증가함에 따라 증가하였으나 감마선 조사에서는 조사하지 않은 대조군과 큰 차이를 나타내지 않았다. Plasmid DNA의 손상 양상은 BSH 존재시 중성자 및 감마선 조사에서 다소 다름을 알 수 있었다.

Hypolipidemic Activities of Dietary Pleurotus ostreatus in Hypercholesterolemic Rats

  • Alam, Nuhu;Yoon, Ki-Nam;Lee, Tae-Soo;Lee, U-Youn
    • Mycobiology
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    • 제39권1호
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    • pp.45-51
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    • 2011
  • This work was conducted to investigate dietary supplementation of oyster mushroom fruiting bodies on biochemical and histological changes in hyper and normocholesterolemic rats. Six-week old female Sprague-Dawley albino rats were divided into three groups of 10 rats each. Feeding a diet containing a 5% powder of Pleurotus ostreatus fruiting bodies to hypercholesterolemic rats reduced plasma total cholesterol, triglyceride, low-density lipoprotein (LDL), total lipid, phospholipids, and LDL/high-density lipoprotein ratio by 30.18, 52.75, 59.62, 34.15, 23.89, and 50%, respectively. Feeding oyster mushrooms also significantly reduced body weight in hypercholesterolemic rats. However, it had no adverse effects on plasma albumin, total bilirubin, direct bilirubin, creatinin, blood urea nitrogen, uric acid, glucose, total protein, calcium, sodium, potassium, chloride, inorganic phosphate, magnesium, or enzyme profiles. Feeding mushroom increased total lipid and cholesterol excretion in feces. The plasma lipoprotein fraction, separated by agarose gel electrophoresis, indicated that P. ostreatus significantly reduced plasma ${\beta}$ and pre-${\beta}$-lipoprotein but increased ${\alpha}$-lipoprotein. A histological study of hepatic cells by conventional hematoxylin-eosin and oil red O staining revealed normal findings for mushroom-fed hypercholesterolemic rats. These results suggest that a 5% P. ostreatus diet supplement provided health benefits by acting on the atherogenic lipid profile in hypercholesterolemic rats.

Apoptosis Induction by Menadione in Human Promyelocytic Leukemia HL-60 Cells

  • Sa, Duck-Jin;Lee, Eun-Jee;Yoo, Byung-Sun
    • Toxicological Research
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    • 제25권3호
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    • pp.113-118
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    • 2009
  • Cell death induced by menadione (vitamin K-3,2-methyl-1,4-naphthoquinone) has been investigated in human promyelocytic leukemia HL-60 cells. Menadione was found to induce both apoptosis and necrosis in HL-60 cells. Low concentration ($1{\sim}$50 ${\mu}$M) of menadione induced apoptotic cell death, which was demonstrated by typical DNA ladder patterns on agarose gel electrophoresis and flow cytometry analysis. In contrast, a high concentration of menadione (100 ${\mu}$M) induced necrotic cell death, which was demonstrated by DNA smear pattern in agarose gel electrophoresis. Necrotic cell death was accompanied with a great reduction of cell viability. Menadione activated caspase-3, as evidenced by both increased protease activity and proteolytic cleavage of 116 kDa poly(ADP-ribose) polymerase (PARP) into 85 kDa cleavage product. Caspase-3 activity was maximum at 50 ${\mu}$M of menadione, and very low at 100 ${\mu}$M of menadione. Taken together, our results showed that menadione induced mixed types of cell death, apoptosis at low concentrations and necrosis at high concentrations in HL-60 cells.

효율적인 Metagenomic Library의 제작 방법 탐구 (Development of an Efficient Procedure for the Construction of Metagenomic Library from Environment Samples)

  • 임동빈
    • 미생물학회지
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    • 제40권4호
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    • pp.359-363
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    • 2004
  • 자연계에 존재하는 미생물의 대부분이 배양이 불가능하다는 것이 밝혀진 이후, 자연계의 시료로부터 직접 유전자를 클로닝하여 유용유전자를 발굴하는 메타제놈(metagenome) 이용 방법이 주목을 받게 되었다. 그러나 실제로 오염이 심한 환경시료로부터 DNA를 추출하여 유용한 메타제놈 라이브러리(metagenomic library)를 제작하기란 쉽지 않은 일이다. 따라서 본 연구에서는 실제 메타제놈 라이브러리를 제작할 때 만나는 기술적 문제점에 대한 해결 방법을 탐구하였다. 메타제놈 라이브러리 제작에는 fosmid vector가 가장 편리하였으며, 성공적인 라이브러리제작에는 fosmid vector에 클로닝이 가능한 40 kbp 크기의 DNA 조각을 얻는 과정이 중요함을 알았다. 여러 실험 조건을 종합적으로 검사한 후 메타제놈 라이브러리 제작에 대한 최적 방법을 제사하였다.