• YAKHAK HOEJI
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• v.29 no.6
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• pp.147-155
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• 1985

이글에서는 예를들어 immunoaffinity chromatography, HPLC및 고압에서 신속하게 분리될수 있게 해주는 새로운 chromatographic media (FPLC)등이 서둘러 개발되고 있는데 본문에섬는 최근에 개발되고 있는 새로운 high resolution 기술을 중심으로 하여 경제성있는 고순도의 rDNA 산물의 회수방법에 대하여 토론하고자 한다. 또한 이들의 정제 방법에 의한 rDNA 산물의 회수에 대한 한계성을 검토하고 최근에 활발히 개발되고 있는 rDNA 조작에 의해 발효산물의 추출및 정제를 용이하게 해줌으로 고수득률과 고순도의 회수를 가능하게 해주는 "upstream processing"기술과 정제 기술의 병용방법에 관하여 집중적으로 토론하고자 한다.

• Proceedings of the KAIS Fall Conference
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• 2007.05a
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• pp.273-275
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• 2007

본 논문은 실리카 코팅 자성 나노입자를 이용한 새로운 DNA의 정제에 관한 것으로 기존의 negative-charged polyanion 형태의 수지 및 실리카 멤브레인 기술을 이용한 제품의 단점인 저속, 고비용의 정제방법을 개선할 수 있는 새로운 정제법을 기술한 것이다.

• Journal of agriculture & life science
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• v.43 no.2
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• pp.31-38
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• 2009

The DNA sequencer is known to be more sensitive for the quality of template DNA, method of purification followed by sequencing reaction, and gel concentration. Therefore, we investigated optimal conditions for template preparation, purification, sequencing reaction, gel concentration, and injection medium. For plasmid prepara- tion, using chloroform instead of phenol improved the average read length from 532 bp to 684 bp. The addition of 2.5% DMSO sequencing PCR reaction resulted in 200 bp longer sequences. Purification using 50 mM EDTA and 0.6 M Sodium acetate(pH 8.0) presented 20 bp longer sequences than that using 50 mM EDTA(pH 8.0) and 0.6 M sodium acetate(pH 5.2). The injection for sequencing analysis using ABI formamide presented 90 bp longer sequences than that of using formamide deionized by resin. Moreover, there were 150 bp more readable sequences in 3.6% PAGE gel than in 4%. Consequently, it was concluded that an average of 700 bp per reaction with 85% accuracy can be obtained by the following optimal conditions: template preparation using chloroform, 2.5% DMSO, 50 mM EDTA and 0.6 M sodium acetate(pH 8.0), ABI formamide and 3.6% gel concentration.

• Journal of Life Science
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• v.16 no.2 s.75
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• pp.240-244
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• 2006

We present a fast and simple protocol for purification of mitochondria, mitochondrial DNA, and RNA from small amounts of tomato leaves. This method uses a high ionic strength medium to isolate mitochondria and extract mitochondrial DNA and RNA from a single preparation and is easily adaptable to other plant species. Mitochondria was confirmed by MitoTracker. The mitochondrial DNA was not contaminated by plastid DNA, was successfully used for PCR. Similarly, the isolated mitochondrial RNA was not contaminated only slightly contaminated (leaves) by plastid RNA. RNA prepared according to our method was acceptable for RT-PCR analysis

• Korean Journal of Fisheries and Aquatic Sciences
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• v.20 no.2
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• pp.136-145
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• 1987

From the sea-urchin, Hemicentrotus pulcherrismus, we have purified by four column chromatographic steps for DNA polymerase $\alpha$ activity. The molecular weight of DNA polymerase u was determined to be around 137,000-138,000 by Sephadex G-200 gel filtration and SDS-polyacrylamide gel electrophoresis. The purified enzyme had the optimal activity at pH 7.4. This enzyme showed to be a function of the metal ion $K^+,\;Na^+$\;and\;Mg^{2+}$ employed as activators, the optimum $K^+$\;or\;Na^+ concentration were 20 mM or 25mM and the optimum $Mg^{2+}$ concentration was 10 mM. The enzyme activity was inhibited by N-ethyl-maleimide, aphidicolin, cytosine $\beta-D-arabinofuranoside$ 5'-triphoshate (ara CTP) and phosphonoacetic acid.

• KSBB Journal
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• v.25 no.2
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• pp.199-204
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• 2010

We generated the feasibility of DNA separation in free-solution using genetically engineered repetitive polypeptides as drag-tags. Two different-sized repetitive polypeptides were designed, expressed in E. coli, and purified. They were conjugated to a fluorescently labeled DNA (100 base), and the electrophoretic mobilities of these conjugate molecules were analyzed on a microchip. The results of these studies indicate that genetically engineered repetitive polypeptide is a prominent candidate for rapid and high-throughput genetic mutation detection, such as SNP analysis.

• Korean Journal of Food Science and Technology
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• v.31 no.1
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• pp.110-114
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• 1999

The present study was conducted to investigate the preparative methods of agarose for gel electrophoresis from agar. Naturally occuring agar consists of two main polysaccharides, the neutral polysaccharide agarose and the acid sulphated polysaccharide agaropectin. The sulphate and carboxyl functions of the agar are accumulated in the agaropectin. The hydrophilic, non-ionogenic, rigid and transparent gel matrix of the agarose was found to be suitable for gel electrophoresis gel filtration and affinity chromatography. Agar was purified by chitosan treatment, cetylpyridinium chloride (CPC) treatment, and polyethylene glycol (PEG) treatment. Yields of agarose purified from agar with chitosan, CPC and PEG were 56.7%, 55.6% and 62.3%. It was proper to treat with chitosan in preparative methods of agarose for gel electrophoresis from agar.

• Korean Journal of Microbiology
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• v.30 no.6
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• pp.466-471
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• 1992

To determine the site at which -1 ribosomal frameshifting occurs within the gag-pro overlap of HTL V-I. DNA fragment corresponding to a portion of the gene overlap was cloned into a SP6 vector. The resultant plasmid harbors the hybrid gene consisting of a synthetic gene encoding 5 amino acids derived from chick prelysozyme including the initiator methionine plus 141 nucleotides of gag-pro overlapping region followed by Staphylococcus aurcus protein A gene fragment. In vitro transcription by SP6 RNA polymerase with this DNA template made an abundant amount of single species mRNA. Cell-free translation programmed with the RNA transcribed in vitro yielded a polypeptide of 21 kDal in size. which could be purified into homogeneity by IgG-Sepharose affinity chromatography. In vitro system described in this study must be useful for rapid purification and sequencing of the Gag-Pro transframe protein. allowing to determine the exact frameshift site on mRNA and to identify the tRNA involved in frameshifting event for the expression of pro gene.

• Journal of Marine Bioscience and Biotechnology
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• v.1 no.2
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• pp.119-125
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• 2006

To understand the comprehensive mechanisms of biological function for insulin-like growth factor-I (IGF-I) in vertebrates, we have investigated the cDNA sequence of this gene in the korean rockfish (Sebastes schlegeli). The mature form of korean rockfish IGF-I was found to be comprised of 67 amino acid residues, showing about a 7 kDa molecular weight. In this study, we used the polymerase chain reaction (PCR) to obtain a korean rockfish IGF-I (KR IGF-I) cDNA fragment, and methods of rapid amplification of cDNA ends (RACE) to obtain a full length of the KR IGF-I sequence. The KR IGF-I encoded for a predicted amino acid sequence showed identities of 93.6 %, 90.7 %, and 85.4 % in comparison with flounder, chinook salmon, and human IGF-I, respectively. To obtain recombinant biologically active polypeptides, korean rockfish B-C-A-D domains were amplified using the PCR, then the isolated cDNA was expressed in the E. coli BL21(DE3). The recombinant KR IGF-I protein biological function was measured by stimulation of [$^3H$] thymidine incorporation, suggesting the cDNA codes for the korean rockfish proIGF-I.

• Applied Biological Chemistry
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• v.45 no.4
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• pp.190-194
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• 2002

Expression of Bacillus subtilis glutamyl-tRNA synthetase (GluRS) in Escherichia coli is lethal for the host, probably because this enzyme misaminoacylates ${tRNA_l}^{Gln}$ with glutamate in vivo. In order to overexpress B. subtilis GluRS, encoded by the gltX gene, in E. coli, this gene was amplified from B. subtilis 168 chromosomal DNA using PCR method and the entire coding region was cloned into a pET11a expression vector so that it was expressed under the control or the T7 Promoter. The resulting recombinant pEBER plasmid was transformed into E. coli Novablue (DE3) bearing the T7 RNA polymerase gene for expression. After IPTG treatment, the overproduced enzyme was purified using ammonium sulfate fractionation, Source Q anion exchange chromatography, Superdex-200 gel filtration, and Mono Q anion exchange chromatography. The purified enzyme yielded 18-fold increase in specific activity over the crude cell extract and its molecular weight was approximately 55 kDa on SDS-PAGE.