• Journal of Plant Biology
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• v.33 no.1
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• pp.73-80
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• 1990

Subcellular distribution of arginase activity was measured in leaves of Canavalia lineata. Both mitochondrial and cytosolic fraction were found to contain the arginase activity. It was noticible that cytosolic fraction contained a substantial amount of arginase activity. Different mobility of arginase from these two fractions was showed on DEAE-Sephacel chromatography and polyacrylamide gel electrophoresis. Also different pI value was showed 6.3 in cytosolic and 6.7, 7.1 in mitochondiral fraction on IEF gel electrophoresis. However, canavaine-dependent-activity (CDA) of arginase in these two fractions were not different. These results indicate that heterogenity of arginase occurs in leaves of C. lineata.

• Journal of the Korean Chemical Society
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• v.48 no.5
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• pp.483-490
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• 2004

We have developed a novel polymerase chain reaction (PCR)-microchip gel electrophoresis (MGE) method based on the sieving gel mixture of commercially available poly(vinylpyrrolidone) (PVP) and hydroxy ethyl cellulose (HEC) for the rapid detection and diagnosis of the orf virus (ORFV) from Korean indigenous goat. After amplification of 594-bp DNA fragment from the B2L gene of ORF virus, the amplicon was analyzed by the MGE separation. The glass microfluidic chip (64 mm total length (36 mm effective length)${\times}$90 ${\mu}$m width${\times}$20 ${\mu}$m depth) allowed the fast detection and diagnosis of ORFV in the mixture of 1.0% PVP ($M_r$ 360,000) and 1.0% HEC ($M_r$250,000) as a sieving matrix with better resolution and reproducibility of DNA fragments. Under the electric field of 277.8 V/cm, the 594-bp DNA was analyzed within 4 min. Compared to traditional slab gel electrophoresis, the PCR-MGE method was twenty times faster and an effective clinical method for the quantitative analysis of ORFV.

• Korean Journal of Microbiology
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• v.20 no.4
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• pp.183-188
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• 1982

The neurotoxin of Clostridium botulinum type B was purified from a liquid culture. The purification steps consist of ammonium sulfate precipitation of whole culture, treatment of Polymin P(0.15%, v/v), gel filtration on Sephadex G-100 at pH5.6 and DEAE-Sephadex charomatography at pH8.0. The procedure recovered 17% of the toxin assayed in the starting culture. The toxin was homogeneous by sodium dodecyl sulfate(SDS)-polyacrylamide gel electrophoresis and had a molecular weight of 163, 000. Subunits of 106, 000 and 56, 000 molecular weight were found when purified toxin was treated with a disulfide-reducing agent and electro phoresed on SDS-polyacrylamide gels.

• Journal of Veterinary Clinics
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• v.11 no.1
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• pp.389-392
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• 1994

To purify transforming growth factor type beta(TGF-$\beta$) in canine platelets, Sephadex G-75 gel filtration and semipreparative HPLC were carried out. The column of $2.0 {\times}120cm$ was used for gel filtration and one inch semipreparative column filled with SP-Toyopeal for HPLC. Electrophoresis and bioassay using African green monkey kidney cell were used for identification of TGF-$\beta$ Crude TGF-$\beta$ of 2.75mg was extracted from 5.2g of the platelets by the treatment of acid/ethanol. In gel filtration of crude TGF-$\beta$, 4 peaks were observed at the detection of spectrophotometer at 280nm. Electrophoresis and bioassay identified the 3rd peak TGF-$\beta$. Linear gradient elution from 0 to 3M NaCl in sornipreparative HPLC showed TGF-$\beta$ at 1.5M NaCl. Gel filtration was less expensive and useful method for the purification of TGF-$\beta$.

• Archives of Pharmacal Research
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• v.25 no.5
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• pp.704-708
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• 2002

A fast and sensitive protein staining method in sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) using both an acidic dye, Coomassie Brilliant Blue R-250 (CBBR) and a basic dye, Neutral Red (NR) is described. It is based on a counter ion-dye staining technique that employs oppositely charged two dyes to form an ion-pair complex. The selective binding of the free dye molecules to proteins in an acidic solution enhances the staining effect of CBBR on protein bands, and also reduces gel background. It is a rapid staining procedure, involving fixing and staining steps with short destaining that are completed in about 1 h. As the result, it showed two to fourfold increase in sensitivity comparing with CBBR staining. The stained protein bands can be visualized at the same time of staining.

• KSBB Journal
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• v.17 no.1
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• pp.106-109
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• 2002

Gel electrophoresis of DNA is a well known technique in molecular biology. This technique is simple, rapid to perform, and capable of adequately separating fragments of DNA. A number of mixtures of DNA fragments ("DNA size markers") are frequently employed in a purpose of extrapolating the sizes or the amount of DNA molecules during gel electrophoresis. DNA size markers are constructed by digesting plasmid DNA, bacteriophage DNA, or recombinant DNA molecules with one or more restriction enzymes. However, liquid suspension containing DNA size marker needs to be kept at a low temperature during storage and shipping. In an attempt to maintain the DNA samples at room temperature for extended period of time, lyophilization of DNA with addition of nuclease inhibitor was studied. Gel loading buffer was also added to the lyophilized DNA to provide additional convenience such that DNA size marker was the "ready-to-use" followed by simply reconstituting with distilled water.

• Journal of Applied Biological Chemistry
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• v.56 no.2
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• pp.95-100
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• 2013

Soil microbes are important integral components of soil ecosystem which have significant and diverse role in organic matter decomposition, nitrogen cycling, and nitrogen fixation. In this study an effective denaturing gradient gel electrophoresis (DGGE) method was employed for paddy soil microbial diversity survey. For optimum paddy soil microbial DNA extraction, different methods such as Lysis buffer, skim milk bead, sodium phosphate buffer, Epicentre Soil Master DNA extraction kit (Epicentre, USA) and Mo Bio Power Soil DNA kit (MO BIO, USA) methods were utilized. Among all the method, using Mo Bio Power Soil kit was most effective. DGGE analysis of Bacteria was carried out at 6% polyacylamide gel and 45-60% denaturing gradient in the optimal conditions. Whereas DGGE analysis of fungi was done at 6% polyacrylamide gel and 45-80% denaturing gradient in the optimal conditions. By applying the above assay, it was found that variation within the microbial community of paddy soil occurs by a factor of time. DGGE assay used in this study through for a variety of soil microbial analysis suggests the potential use of this method.

• Journal of Microbiology and Biotechnology
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• v.10 no.3
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• pp.405-409
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• 2000

We have studied the genome of an obligately commensal thermophile, Symbiobacterium toebii. The chromosome was extracted from pure cultures of S. toebii recently established. Total DNA of S. toebii was resolved by pulsed-field gel electrophoresis (PFGE) into discrete numbers of fragments by digenstion with the endonuclease SspI, SpeI, XbaI, and HpaI. Estimated sizes of fragments produced by the four enzymes and their sum consistently yielded a total genome size of 2.8 Mb. Because restriction endonucleases NotI and SwaI, recognizing 8 bp, released too many fragments, these enzymes could not be used for the estimation of the genome size. Considering no mobility of undigested genome under PFGE, the genome of S. toebii appears to be circular. The presence of extrachromosomal DNA in S. toebii was excluded by the results of the conventional 1% agarose gel electrophoresis and the field inversion gel electrophoresis of undigested S. toebii DNA.

• Environmental Mutagens and Carcinogens
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• v.21 no.2
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• pp.89-94
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• 2001

Recently, single cell gel electrophoresis, also known as comet assay, is widely used for the detection and measurement of DNA strand breaks in vitro and in vivo in many toxicological fields such as radiation exposure, human monitoring and toxicity evaluation. As well defined, comet assay is a sensitive, rapid and visual method for the detection of DNA strand breaks in individual cells. Briefly, a small number of damaged cells suspended in a thin agarose gel on a microscope slide were lysed, unwinded, electrophoresed, and stained with a fluorescent DNA binding dye. The electric current pulled the charged DNA from the nucleus such that relaxed and broken DNA fragments migrated further. The resulting images which were subsequently named for their appearance as comets, were measured to determine the extent of DNA damages. However, some variations could be occurred in procedures, laboratories's conditions and kind of cells used. Hence, to overcome and to harmonize these matters in comet assay, International Workshop on Genotoxicity Test Procedure (IWGTP) was held with several topics including comet assay at Washington D.C. on March, 1999. In spite of some consensus in procedures and conditions in IWGTP, there are some problems still remained to be solved. In this respect, we attempted to set the practical optimal conditions in the experimental procedures such as lysis, unwinding, electrophoresis and neutralization conditions and so on. First of all, we determined optimal lysis and unwinding time by using 150 $\mu$M methyl methanesulfonate (MMS) which is usually used concentration. And then, we determined optimal positive control concentrations of benzo(a)pyrene (BaP) and MMS in the presence and absence of S9 metabolic activation system, respectively.

• Korean Journal of Microbiology
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• v.30 no.3
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• pp.160-163
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• 1992

Killer toxin from killer yeast fusant FWKS 260 developed by protoplast fusion between the wild killer yeast and alcohol-fermenting yeast was purified by ammonium sulfate fractionation. Amicon PM I0 concentration. Sephadex G-200 and Scphadcx G-75 column chromatography. The purified killer toxin showed a single band by SIX-polyacvlamide gel electrophoresis. The protein part of killer toxin was active site. which was found by treating the proteolytic enzyme such as pronase E and pepsin to killer toxin. The killer toxin was stable at pH 2.0-5.0 and 20$^{\circ}$C. but inactivated with increasing temperature. The molecular weight was determined to be approximately 13.000 according to the results obtained from the SDS-polyacrylamide gel electrophoresis. It was confirmed that the purified killer toxin is glycoprotein by showing a red single band after st'tining with Schiffs reagent.