• Parasites, Hosts and Diseases
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• v.46 no.3
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• pp.157-164
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• 2008

Three Acanthamoeba isolates (KA/E9, KA/E17, and KA/E23) from patients with keratitis were identified as Acanthamoeba triangularis by analysis of their molecular characteristics, a species not previously recognized to be a corneal pathogen. Epidemiologic significance of A. triangularis as a keratopathogen in Korea has been discussed. Morphologic features of Acanthamoeba cysts were examined under a microscope with differential interference contrast (DIC) optics. Mitochondrial DNA (mtDNA) of the ocular isolates KA/E9, KA/E17, and KA/E23 were digested with restriction enzymes, and the restriction patterns were compared with those of reference strains. Complete nuclear 188 and mitochondrial (mt) 16S rDNA sequences were subjected to phylogenetic analysis and species identification. mtDNA RFLP of 3 isolates showed very similar patterns to those of SH621, the type strain of A. triangularis. 16S and 18S rDNA sequence analysis confirmed 3 isolates to be A. triangularis. 18S rDNA sequence differences of the isolates were 1.3% to 1.6% and those of 16S rDNA, 0.4% to 0.9% from A. triangularis SH621. To the best of our knowledge, this is the first report, confirmed by 18S and 16S rDNA sequence analysis, of keratitis caused by A. triangularis of which the type strain was isolated from human feces. Six isolates of A. triangularis had been reported from contaminated contact lens cases in southeastern Korea.

• Transactions of the Korean Society of Mechanical Engineers A
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• v.28 no.10
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• pp.1598-1602
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• 2004

In this paper, we report a microthermostat using PDMS (poly-dimethylsiloxane) and glass. This PDMS/glass chip is able to maintain constant temperature that is necessary for restriction enzyme reaction. Since PDMS is the low-cost and the mass-producible material and has very good biochemical compatibility, PDMS chip has more benefit than general Si chip. Heater was made of Au wiring patterned on Pyrex glass. A reaction chamber has a capacity of about 3 ${mu}ell$. We performed a restriction enzyme reaction by using the fabricated microthermostat and conventional method. Then, with the electrophoresis, we made a comparison between the result from the micro reactor and the one from conventional method.

• Journal of Life Science
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• v.22 no.2
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• pp.245-250
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• 2012

The 16S rDNA - RFLP types for six Vibrio species (V. fluvialis, V. proteolyticus, V. vulnificus, V. mimicus) including two core group members, V. alginolyticus and V. parahaemolyticu s, and Grimontia (Vibrio) hollisae were determined using PCR-RFLP analysis. Six tetrameric restriction enzymes (Alu I, Cfo I, Dde I, Hae III, Msp I, and Rsa I) were selected for RFLP analysis. V. alginolyticus, V. parahaemolyticus, and V. proteolyticus showed the same RFLP pattern following digestion with four of the six used restriction enzymes: CfoI, DdeI, MspI, and RsaI. Various restriction enzyme combinations generated digests recognizable as distinct RFLP types for each of the assayed Vibrio species. In particular, AluI single digestion produced species specific band patterns that enabled the differentiation between these Vibrio species. Dendrogram based on restriction patterns showed that two Vibrio core group members, V. alginolyticus and V. parahaemolyticus were closely related having a similarity over 90%. Although the observed RFLP pattern for Grimontia hollisae shared several common bands with other Vibrio spp., G. hollisae results were still clearly distinct from Vibrio spp. RFLP types for all restriction enzymes tested. If restriction enzymes are aptly selected, PCR-RFLP analysis is still a rapid and effective tool for differentiating Vibrio species.

• Korean Journal of Microbiology
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• v.24 no.4
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• pp.357-363
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• 1986

Synthesis of threonine and methionine in yeast, Saccharomyces cerevisiae shares a common pathway from aspartate via homoserine. HOM6 gene encodes homoserine dehydrogenase (HSDH) which catalyzes the inter-conversion of beta-aspartate semialdehyde and homoserine. The level of HSDH is under methionine specific control. A recombinant plasmid (pEK1: 13.3kb), containing HOM6 gene, has been isolated and cloned into E. coli by complenemtary transformation of a homoserine auxotrophic yeast strain M-20-20D (hom6, trp1, ura3) to a prototrophic M20-20D/pEK1, using a library of yeast genomic DNA fragments in a yeast centromeric plasmid, YCp50(8.0kb). Isolation of HOM6has been primarily confirmed by retransformation of the original yeast strain M20-20D, using the recombinant plasmid DNA which was extracted from M20-20D/pEK1 and subsequently amplified in E. coli. Eleven cleavage sites in the insery (5.3kb) have been localized through fragment analysis for 8 restriction endonucleases; Bgl II(2 site), Bgl II(1), Cla I(3), Eco RI(1), Hind III(2), Kpn I (1), Pvu II(1) and Xho I(1).

• Journal of Periodontal and Implant Science
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• v.25 no.3
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• pp.679-693
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• 1995

The present study has been performed to see the intrafamilial transmission of periodontopathic organism Actinobacillus actinomycetemcomitans in Koreans having various froms of periodontal diseases. 17 clinical isolates from 8 periodontal patients and 20m clinical isolates from their 8 family members were grown anaerobically for the serotyping and the extraction of genomic DNA. The DNA was digested with restriction endonucleases (EcoRI+HindIII) and plasmid pAA 2097(kindly provided by Dr. DiRienzo, Univ. of Pennsylvania) including 4.7kb-size randlomly clone probe for restriction length pleomorphism analysis(RFLP). RFLP patterns of reference serotypes a, b, c, d, and e were used as the genotypes A, B, C, D, and E, respectively for comparison of genotypes of clinical isolates. 28 out of total 37 clinical isoltes belonged to either one of 5 basic gentotypes and 9 remaining isolats did not fall into any types, and hence were designate as non-type(NT). Genotype C were the most frequently found one(35.1%) and genotype B has not isolated. Intrafamilial transmission of bacteria between spouses, brothers and sisters, and parents and their offsprings, resepctively could well be demonstrated by comparing RFLP patterns. There were not any specific genotypes which showed predominance over others in terms of transmission.

• BMB Reports
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• v.29 no.1
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• pp.52-57
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• 1996

A specific DNA fragment from Korean radish (Raphanus sativus L.) was amplified by performing PCR with oligonucleotide primers which correspond to the highly conserved regions of plant peroxidases. The size of the PCR product was ca. 400 bp, as expected from the known plant peroxidase genes. Comparison of the nucleotide and deduced amino acid sequences of the PCR product to those of other plant peroxidase-encoding genes revealed that the amplified fragment corresponded to the highly conserved region I and III of plant peroxidases. By screening a genomic library of Korean radish using the amplified fragment as a probe, two positive clones, named prxK1 and prxK2, were isolated. Restriction mapping studies indicated that the 5.2 kb Sail fragment of the prxK1 clone and the 4.0 kb EcoRI fragment of the prxK2 clone encode separate isoperoxidase genes. Analyses of the promoter region of the prxK1 clone shows that putative CAAT box, CMT box, and TGA1b binding sequence (5' TGACGT) are present 718 bp upstream from the start codon.

• Korean Journal of Microbiology
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• v.36 no.4
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• pp.249-253
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• 2000

The genetic diversity among Korean cyanobacteria was assessed by restriction fragment length polymorphism(RFLP) analysis of PCR products from the phycocyanin locus. Strains of all the genera tested were successfully amplified, and the size of amplified fragments was approximately 700bp. The restriction patterns generated by AluI, MspI, and HaeIII were conserved for strains within each of genera studied and were specific to the genus level. Intrageneric delineation of strains was revealed by the enzyme, CfoI for members of genera Anabeana and Synechocystis. Phenogram derived from the different RFLP patterns revealed a coherent cluster among Anabeana, Chlorogloea, and Synechocystis strains. PC-RFLP methods provided useful tools for classification of cyanobacteria.

• Korean Journal of Clinical Laboratory Science
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• v.38 no.3
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• pp.158-165
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• 2006

Although Mycobacterium tuberculosis complex strains remain responsible for the majority of diseases caused by mycobacterial infections worldwide, the increase in HIV infections has allowed for the emergence of other non-tuberculous mycobacteria as clinically significant pathogens. However, Mycobacterium species has a long period of incubation, and requires serious biochemical tests such as niacin, catalase, and nitrate test that are often tedious. The development of rapid and accurate diagnostics can aid in the early diagnosis of disease caused by Mycobacterium. The current DNA amplification and hybridization methods that have been developed target several genes for the detection of mycobacterial species such as hps65, 16S rDNA, rpoB, and dnaj. These methods produce rapid and accurate results. In this study, PCR-restriction fragment length polymorphism analysis(PCR-RFLP) based on the region of the rpoB gene was used to verify the identification of non-tuburculosis Mycobacterium species. A total of 8 mycobacterial reference strains and 13 clinical isolates were digested with restriction enzymes such as Msp I in this study. The results of using this process clearly demonstrated that all 13 specimens were identified by rpoB gene PRA method. The PCR-RFLP method based on the rpoB gene is a simple, rapid, and accurate test for the identification of Mycobacterium.

• Korean Journal of Clinical Laboratory Science
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• v.37 no.3
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• pp.173-177
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• 2005

YMDD motif mutants of the hepatitis B virus(HBV) emerge in chronic hepatitis B patients after prolonged lamivudine treatment. HBV DNA breakthrough may be accompained by the emergence of YMDD mutants. We compared the performance of the sequencing method with that of RFMP method in chronic hepatitis B patients who had suffered the HBV DNA breakthrough after lamivudine treatment. Both sequencing and RFMP methods were used to detect YMDD variants in 20 chronic hepatitis B patients. YMDD mutants were detected in 17 samples (85.0%) by both methods. Among them, no mutants were detected in two samples(10.0%), while they were detected in the other sample(5.0%) with the RFMP method. The concordance rate between both methods was 95.0%. There was inconsistency in one sample showing mutants detected by the RFMP method, but not by sequencing method. In the sequencing method, the mutants was detected in the major type virus, but not in the minor type virus. However, both sequencing and RFMP methods were highly concordant except in one sample, so it is suggested that both methods are useful to detect YMDD mutants of chronic hepatitis B patients.

• Journal of the Korean Society of Food Science and Nutrition
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• v.27 no.6
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• pp.1166-1172
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• 1998

A method for the isolation and purification of DNA from a red algae, Porphyra was innovated. The innovation of the method consists mainly of three steps that include sodium acetate treatment, chloroform extraction, and 0.2 volume isopropanol precipitation step. The sodium acetate treatment was designed to remove polysaccharide contamination, and the isopropanol step to remove proteins and salts contaminents. Genomic DNA,s of several species(for example, P. tenera, P. yezoensis, P. seriata, and P. pseudolinearis) was successfully isolated by the innovated method. The amount of DNA purified from one g of sample material with the innovated method was 53 g in average. The resulting DNA was characterized to include high molecular weight and showed no nuclease activity. The DNA was pure enough to be digested directly by various restriction enzymes without any difficulties. Porphyra DNA was pure enough and adequate for amplification reaction through the polymerase chain reaction (small nuclear rDNA PCR amplification).