• BMB Reports
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• v.38 no.4
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• pp.491-499
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• 2005

DNA-based molecular markers for differentiation of five penaeid shrimps (Penaeus monodon, P. semisulcatus, Feneropenaeus merguiensis, Litopenaeus vannamei and Marsupenaeus japonicus) were developed based on polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) and single-stranded conformation polymorphism (SSCP) of 16S ribosomal (r) DNA. Differentiation of P. monodon, P. semisulcatus and L. vannamei can be unambiguously carried out by PCR-RFLP of 16S $rDNA_{560}$ whereas P. semisulcatus and M. japonicus shared a BABB mitotype. These shrimps were successfully discriminated by SSCP analysis of 16S $rDNA_{560}$. Nevertheless, the amplification success for L. vannamei and F. merguiensis was not consistent when tested against larger sample sizes. As a result, 16S $rDNA_{560}$ of an individual representing the most common mitotype of each species was cloned and sequenced. The new primer pair was designed and tested against the large sample sizes (312 bp product, N = 185). The amplification success was consistent across all species. PCR-RFLP of 16S $rDNA_{312}$ was as effective as that of 16S $rDNA_{560}$. Differentiation of all shrimp species were successfully carried out by SSCP analysis.

• Journal of Ecology and Environment
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• v.31 no.4
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• pp.309-316
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• 2008

To develop a monitoring method for soil microbial communities in rice paddy fields, we used terminal restriction fragment length polymorphism (T-RFLP) to compare soil bacterial community structure in rice paddy fields experiencing different management practices: organic practices, conventional practices without a winter barley rotation, and conventional practices with a winter barley rotation. Restriction fragment length profiles from soils farmed using organic practices showed very different patterns from those from conventional practices with and without barley rotation. In principal component analyses, restriction fragment profiles in organic practice samples were clearly separated from those in conventional practice samples, while principal component analysis did not show a clear separation for soils farmed using conventional practices with and without barley rotation. The cluster analysis showed that the bacterial species compositions of soils under organic practices were significantly different from those under conventional practices at the 95% level, but soils under conventional practice with and without barley rotation did not significantly differ. Although the loadings from principal component analyses and the Ribosomal DNA Project II databases suggested candidate species important for soils under organic farming practices, it was very difficult to get detailed bacterial species information from terminal restriction fragment length polymorphism. Rank-abundance diagrams and diversity indices showed that restriction fragment peaks under organic farming showed high Pielou's Evenness Index and the reciprocal of Simpson Index suggesting high bacterial diversity in organically farmed soils.

• Korean Journal of Microbiology
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• v.25 no.1
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• pp.40-45
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• 1987

The gene for the Bdi I modification enzyme, which is one of Bdi I restriction-modification system, fromBrevibacterium divaricatum FERM 5948 was cloned and expressed in E. coli. For cloning of the Bdi I methylase gene, we have initially used three cloning site(EcoRI, BamHI and Sal I) of plasmid vector pBR 322 and adopted the retransformation method after Bdi I restriction endonuclease cleavage. Selection of transformants carrying the gene was based on the resistance of the modified plasmid encoding the enzyme to cleavage by Bdi I restriction enzyme, and the recombinant plasmid pBDIM 116 containing 5.6kb EcoRI insery was proved to carry the gene. Crude cell extracts prepared from strains carrying the plasmid pBDIM 116 contained an S-adenosylmethionine-dependent methyltransferase activity specific for the Bdi I recognition site, ATCGAT. The restriction map was constructed with 11 restriction enzyme, and the Bdi I restriction-modification system was also discussed.

• Korean Journal of Head & Neck Oncology
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• v.12 no.2
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• pp.169-176
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• 1996

Tuberculous cervical lymphadenitis is still an important cause of neck mass in Korea. Tuberculosis is an important differential diagnosis in patients of cervical lymphadenopathy. Rapid and sensitive test for the diagnosis of tuberculosis is essential for the approapiate treatment. Up to now, conventional diagnostic methods for M. tuberculosis were acid-fast bacilli(AFB) stain and culture of M. tuberculosis. The direct microscopic examination of AFB by Ziehl-Neelsen stain is rapid, but often negative. The culture for M. tuberculosis is time-consuming, taking 4 to 8 weeks. Recently various methods to detect Mycobacterial DNA, including PCR and restriction fragment length polymorphism(RFLP) analysis have been reported. Here we represent a simple method for the confirmation of M. tuberculosis and exclusion of the other Mycobacterial species by RFLP analysis and silver staining of polyacrylamide gel electrophoresis after nested PCR for a repetitive DNA sequence(IS986) specific for M. tuberculosis from fresh or paraffin-embedded biopsy specimens. This result leads us to conclude that this method is simple, rapid and possibly applicable to confirm M. tuberculosis and rule out the other Mycobacteria species from the clinical specimens in the clinical laboratories.

• Korean Journal of Microbiology
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• v.48 no.4
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• pp.262-269
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• 2012

Seasonal differences of the cultivable bacterial communities associated with the marine sponge, Hymeniacidon sinapium, between spring and summer were analyzed through the Amplified Ribosomal DNA Restriction Analysis (ARDRA). For the cultivation of the bacterial isolates, modified Zobell and MA media were used. The 16S rDNA of individual strains were amplified and fragmented by using two restriction enzymes, HaeIII and MspI. As a result, 23 ARDRA types from the spring sponge and 28 types from the summer sponge were obtained. The partial sequencing result of 1 to 3 selected strains from each types showed over 94% similarities with the known species from the public database. The bacterial communities from the sponge, captured on spring, contained 4 phyla: Actinobacteria, Alphaproteobacteria, Gammaproteobacteria, and Firmicutes. There were 5 phyla observed from the bacterial communities associated with the sponge, captured on summer: Alphaproteobacteria, Betaproteobacteria, Gammaproteobacteria, Firmicutes, and Bacteroidetes. Gammaproteobacteria was predominant group in both spring and summer, accounted for 33.8% of total in spring and 67.4% in summer, showed increase pattern on summer. Because Firmicutes and Actinobacteria participated in 30.2% and 8.3% of the spring sponge while they represented only 6.9% and 0% of the summer sponge, both bacterial groups showed decrease drift on summer. Betaproteobacteria (4.7%) and Bacteroidetes (4.7%) were only observed on the sponge captured on summer. On the sponge, Hymeniacidon sinapium, more diverse bacterial communities were shown on summer than on spring, and even from the same sponge, there were seasonal differences.

• Korean Journal of Microbiology
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• v.40 no.3
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• pp.189-198
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• 2004

T-RFLP analysis and clone sequencing analysis based on bacterial 16S rDNA were conducted to assess bacterial community structure and diversity in two layers (0-1cm, 6-7cm depth) of the sediment from Janghwari intertidal flat in Ganghwa Island. The results of T-RFLP (terminal-restriction fragment length polymorphism) analysis using restriction enzyme HhaI showed that the T-RFs of various size ($60{\pm}2$) bp-($667{\pm}2$) bp) appeared evenly at the surface sediments but two T-RFs with 60(${\pm}2$)bp and 93 (${\pm}2$)bp predominated at 6-7cm depth. Analysis of partial sequences for 172 clones revealed that 98% of the clones were not matched with the sequences of cultured bacteria strains in the GenBank (${\geq}similarity$ 98%), and approximately 86% of them were classified as different phylotypes. Most clones belonged to $\alpha$-, $\gamma$-, and $\delta$-Proteobacteria, Acidobacteria/Holophaga and green nonsulfur bacteria group. Proteobacteria group occupied the highest proportion in both layers (69% at 0-1cm depth and 46% at 6-7cm depth). $\gamma$-Proteobacteria and $\delta$-Proteobacteria that are associated with oxidation and reduction of sulfur compounds were appeared to be dominant, and comprised 21.5% and 15.7% of total clones, respectively. Overall results indicated that extremely diverse bacterial groups were inhabiting in the sediment of Ganghwa intertidal flat, and bacterial communities associated with the behaviour of sulfur seemed to playa significant role in the biogeochemical environment in this anoxic sediment.

• Korean Journal of Microbiology
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• v.42 no.2
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• pp.103-110
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• 2006

T-RFLP analysis and clone sequencing analysis based on bacterial 16S rDNA were conducted to assess bacterial community structure and diversity in Zoysia japonica soil treated with liquid fertilizer containing amino acids(LFcAA) after spray with herbicide. The results of T-RFLP (terminal restriction fragment length poly-morphism) analysis using restriction enzyme Hae III showed that the T-RFs of various size appeared evenly in the 32 clones of KD3 and 38 clones of KD4 respectively that had been treated with liquid fertilizer containing amino acid(LFcAA) compared to 23 clones of KD2 hat had not been treated with LFcAA. The microbial com- munity structure in KD2 appeared less diverse than those in KD3 and KD4. Analysis of partial sequences for 110 clones from KDI (control), KD2 (non-treated), KD3 (LFcAA 1X), KD4 (LFcAA 2X), respectively, revealed that most bacteria were related with uncultured bacteria in a 16S rDNA sequence similarity range of 91-99% through blast search. Otherwise, the other clones were members of proteobacteria, Acidobacteria, Act-inobacteria, Sphingobacteria and Planctomyces groups. Especially in KD4, members of Alpha Proteobacteria, Rhizobiales, Sphigomonadales, Caulobacterales, Gamma Proteobacteria, the genus Pseudomonas, Betapro-teobacteria, Nitrosomonadales and genus Nitrosospira appeared to be dominant. In addition, Acidobacteria group, Actinobacteria group, Planctomycetacia and Sphingobacteria were also shown. The microbial com-munity structure in Z. japonica soil sprayed with herbicide was affected by LFcAA.

• Journal of Life Science
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• v.8 no.3
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• pp.348-351
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• 1998

Common DNA fragments of the ${\beta}$-globin gene were observed from six races of the adult common carp: Hybrid-Yamato, Japanese wild type, Mirror, Suwa-Yamato, Scale German, and Saku-Yamato. Chromosomal DNAs isolated from the above six races were digested with restriction endonucleased EcoRI and PstI. The digested fragments were transferred onto nitrocellulose filter and hybridized with a probe of carp ${\beta}$-globin cDNA. Molecular sizes of the hybridized DNA fragments digested with EcoRI were 3.6Kb(Kilo base), 4.3Kb and 15Kb in Hybrid-Yamato, Japanese wild type, Mirror, Scale German and Saku-Yamato carp DNAs. In Scale German and Saku-Yamato carp DNAs, two and one more hybridized DNA fragments were observed, respectively. Molecular sizes of the hybridized DNA fragments digested with PstI were 2.2Kb, 6.5Kb, 7.8Kb and 9.2Kb in Hybrid-Yamato, 2.2Kb, 6.5Kb and 9.2Kb in Japanese wild type, 2.2Kb, 6.5Kb, 7.8Kb, and 13Kb in Mirror, 2,2Kb, 5,5Kb, 6.5Kb, 7.8Kb, 9.2Kb and13Kb in Scale German, and 2.2Kb, 5.5Kb, 6.5Kb, 9.2Kb and Saku-Yamato carp DNA. Therefore, depending on carps, three to six DNA fragments were hybridized with ${\beta}$-globin gene probe. Thus it indicated polymorphysm in the globin gene family of carp.

• Biomedical Science Letters
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• v.11 no.2
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• pp.227-235
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• 2005

It has been reported that endurance performance is influenced by various environmental and genetic factors. In view of an important role of human mitochondrial DNA (mtDNA) as a candidate for endurance performance, this study focused on the relationships between $VO_{2max}$ value as a measure of endurance performance or other associated phenotypes and four mtDNA restriction fragment length polymorphisms (RFLPs) (Bam HI, Hinc II1, Hinc II2 and Nci I) in the NADH dehydrogenase subunit 5 and one (Kpn I) in the D-loop region of mtDNA. MtDNA was purified from buffy coat in human peripheral blood, and PCR-RFLP analysis was performed to estimate the allele frequencies of each polymorphism in the mtDNA. There were no significant differences in allele distributions of all polymorphisms studied between male athletes and controls, respectively (P>0.05). However, the Kpn I polymorphism was significantly associated with diastolic blood pressure level in male athletes, respectively (P<0.05). Therefore, our results suggest that this polymorphism might be one of the factors modifying inter-individual difference in cardiovascular risk. Further studies using larger sample size will be required to generalize these results from the study described herein.

• Research in Plant Disease
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• v.17 no.1
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• pp.99-101
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• 2011

The ascomycetous fungus Taphrina wiesneri, the pathogen of cherry witches' broom, is highly pathogenic to Prunus yedoensis, the most widely planted cherry trees in Korea as park and roadside trees. A collection of 13 strains of the pathogen in Korea and Japan was characterized by 18S rDNA gene sequence and restriction fragment length polymorphism (RFLP) analysis. In cluster analysis based on 18S rDNA gene sequence the strains were divided into 2 clusters. In RFLP analysis of the rDNA-IGS region using HhaI, the strains were separated into four patterns, B, C, D and G, of which pattern G was new.