• Journal of Genetic Medicine
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• v.5 no.1
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• pp.15-20
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• 2008

Purpose : Large exon deletions in the DMD gene are found in about 60% of DMD/BMD patients. Multiplex PCR has been employed to detect the deletion mutation, which frequently generates noise PCR products due to the presence of multiple primers in a single reaction as well as the stringency of PCR conditions. This often leads to a false-negative or false-positive result. To address this problematic issue, we introduced the dual primer oligonucleotide (DPO) system. DPO contains two separate priming regions joined by a polydeoxyinosine linker that results in high PCR specificity even under suboptimal PCR conditions. Methods : We tested 50 healthy male controls, 50 patients with deletion mutation as deletion-positive patient controls, and 20 patients with no deletions as deletion-negative patient controls using DPO-multiplex PCR. Both the presence and extent of deletion were verified by simplex PCR spanning the promoter region (PM) and 18 exons including exons 3, 4, 6, 8, 12, 13, 17, 19, 43-48, 50-52, and 60 in all 120 controls. Results : DPO-multiplex PCR showed 100% sensitivity and specificity for the detection a deletion. However, it showed 97.1% sensitivity and 100% specificity for determining the extent of deletions. Conclusion : The DPO-multiplex PCR method is a useful molecular test to detect large deletions of DMD for the diagnosis of patients with DMD/BMD because it is easy to perform, fast, and cost-effective and has excellent sensitivity and specificity.

• The Plant Pathology Journal
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• v.20 no.3
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• pp.200-205
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• 2004

The partial nucleotide sequences of the genomic dsRNA mycovirus infecting Pleurotus ostreatus isolates ASI2223 and Suhan were determined and compared with those of mycoviruses belonging to partitiviruses and totiviruses. Partial nucleotide sequences of the purified dsRNA from ASI2223 and Suhan showed RNA-dependent RNA polymerase sequences that are closely related to those of partitiviruses, including Fusarium poae virus 1, Fusarium solani virus, Rhizoctoniasolani virus, Discula destructiva virus 2, and Oyster mushroom isometric virus 2. Specific primers were designed for RT-PCR detection of dsRNA viruses from the P. ostreatus isolate ASI2223 and Suhan. Two virus specific primer sets were found to specifically detect each virus among six sets of designed oligonucleotide primers. Collectively, these results suggest that dsRNA mycoviruses from P. ostreatus isolates ASI2223 and Suhan belong to the family Partitiviridae, although, they are not the same virus species. Our results also suggest that these virus-specific primer sets can be employed for the specific detection of each viral sequence in infected tissues.

• KOREAN JOURNAL OF CROP SCIENCE
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• v.52 no.1
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• pp.51-54
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• 2007

Soybean has a morphological type with a broadened and flattened stem. Fasciation has been suggested as a new gene for soybean research. SSR marker linked to the $\Large f$ locus that controls fasciation phenotype has not identified within 10 cM. A mapping population consisting of 94 $F_2$ progenies was derived from a cross between wild type Clark (FF) and fasciation mutant C32 (${\Large f}{\Large f}$). The phenotype of $F_2$ individual plants was recorded at R2 and R3 growth stage from field. One-thousand 10-mer oligonucleotide RAPD primers and 29 SSR primers selected from the D1b+W of the soybean molecular linkage map were used. A genetic map was constructed from the segregating 35 RAPD, four SSR markers and one phenotypic(wild type/fasciation) marker. The segregation ratios of 3 : 1 observed in the $F_2$ population and the Chi-square values strongly suggest that the fasciation trait is controlled by a single recessive gene. Satt537 marker was linked to $\Large f$ locus at a distance of 9.6 cM. Assignment of the $\Large f$ locus to linkage group D1b+W and identification of markers can be used as an initial step for fine mapping of the $\Large f$ gene.

• Parasites, Hosts and Diseases
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• v.31 no.3
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• pp.285-292
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• 1993

C. sinensis total RMh was containing large amount of 185 rRNA but little 285 rRNA. The size of the double-stranded cDNA synthesized from poly $(A)^{+}$ mRNA was 0.4-4.2 kb long with tapering unto 9.5 kb. Degenerated oligonucleotides (as 2 sense and 3 antisense Primers) were designed on the conserved regions of the known tropomyosin amino acid sequences. From one out of the PCR amplifications using total CDNA and matrix of primers, a specific gene product, 580 bp in size, was produced. Upon Southern hybridization of the PCR products with Schistosomn mnnsoni tropomyosin (SMTM) CDNA, only one signal appeared at the band of 580 bp product. This 580 bp product was considered to encode C. sinensis tropomyosin (CSTM) and cloned in pGEM-3Zf(-) for DNA sequencing. CSTM cDNA was 575 bp containing one open reading frame of 191 predicted amino acids, which revealed 86.3% homology with SMTM and 51.1% with rrichostronsylur coeubnlormis tropomyosin. CSTM cDNA obtained will serve as a probe in the studies of molecular cloning of CSTM.

• Parasites, Hosts and Diseases
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• v.45 no.1 s.141
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• pp.69-74
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• 2007

Trichostrongylus eggs observed in cellophane-thick smears are difficult, in practice, to distinguish from hookworm eggs. In order to overcome these limitations, a molecular approach was conducted. A Trichostrongylus colubriformis adult worm was obtained from a human in Laos, which was identified morphologically. ITS-1 sequence of this worm was determined, and found to be most similar with that of T. colubriformis among the Trichostrongylus spp. reported so far. Then, this sequence was compared with those of human hookworm species, Ancylostoma duodenale and Necator americanus, and species-specific oligonucleotide primers were designed. Polymerase chain reaction(PCR) using these primers evidenced specifically amplified PCR products of Trichostrongylus sp., A. duodenale and N. americanus from the eggs of each(520 bp, 690 bp, and 870 bp, respectively). A species-specific PCR technique can be developed in order to study the epidemiology of Trichostrongylus spp. and hookworms in endemic areas.

• KOREAN JOURNAL OF CROP SCIENCE
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• v.51 no.2
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• pp.169-172
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• 2006

Leaflet number of soybean controlled by Lf2 locus is the important trait in photosynthesis and plant type. The objective of this research was to identity molecular markers linked to the lf2 locus. A total of $115F_2$ plants were derived from a cross between normal three-leaflet type Sinpaldalkong (Lf2Lf2) and seven-leaflet mutant type T255 (lf2lf2). All leaflet counts of parents and $F_2$ individual plants were made in the field on fully expanded leaves on the main stem when terminal growth of the main stem had ceased. One-thousand 10-mer oligonucleotide RAPD primers and 664 SSR primers were used. The segregation ratios of 3 : 1 were observed in the $F_2$ population and the Chi-square values strongly suggested that the seven-leaflet was controlled by a single recessive gene. A genetic map was constructed from the 15 segregating markers (9 RAPDs, 5 SSRs, 1 lf2 locus). OPAD03 and OPAI13 RAPD markers were linked to the lf2 locus that controlled seven-leaflet type at a distance of 20.5 and 23.5 cM, respectively. Molecular markers identified in this study linked with lf2 locus will be helpful to locate lf2 locus on the public soybean molecular linkage map and would be useful for tagging the lf2 locus that controls seven-leaflet trait.

• Korean Journal of Medicinal Crop Science
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• v.8 no.3
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• pp.243-249
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• 2000

Analysis of random amplified polymorphic DNAs(RAPDs) and internal morphological features were performed using three species of medicinal plants in the genus of Angelica(A. gigas Nakai, A. sinensis(Oliv.) Diels., A. acutiloba Kitagawa) to distinguish between these three species. Fifty decarmer oligonucleotide primers were screened for the RAPDs of the herbal plant species. Five primers generated distinct RAPD markers specific to the species of Angelica, In analysis of the degree of similarity, A. sinensis(Oliv.) Diels is more closely related to A. acutiloba Kitagawa than to A. gigas Nakai. Furthermore, we proved the usefulness of RAPD analysis for the discrimination of the species using dry roots and commercial plant materials. In internal morphology of three species, A. sinensis(Oliv.) Diels seemed to be more specialized in systemic than A. acutiloba Kitagawa and A. gigas Nakai

• The Journal of Korean Society of Virology
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• v.27 no.2
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• pp.217-225
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• 1997

PCR amplication using the primers for gag, pol and env genes in BLV (bovine leukemia virus) proviral DNA and syncytium assay were carried out for the Korean native goats experimentally infected with bovine leukemia virus to investigate pathogenesis of BLV in the goats, and to establish a model animal for BLV infection. The oligonucleotide primers used in PCR revealed very high specificity. The minimal amount of FLK-BLV cellular chromosomal DNA to detect the integrated BLV proviral DNA was 10 ng. The peripheral blood lymphocytes from the goat infected with BLV were examined at regular intervals by PCR amplification and syncytium assay. Pol or gag genes were detected in none of three infected goats at the 1st week post-infection (p.i.). At the 4th week p.i., one of three goats showed the amplified gag gene. Thereafter detection rates for the genes were increased, indicating that the BLV proviral genes were integrated in all of the lymphocytes from three goats, at the 16th weeks p.i., when it was evident in syncytium assay that the lymphocytes from all of three goats were infested with infective BLV. Investigating the tissues from the necropsied goats at the 8th month p.i., the amplified BLV proviral genes and infective BLV were detected in all of the peripheral lymphocytes from three infected-goats. Among various tissues examined, the amplified BLV proviral genes were observed in spleen and superficial cervical, mandibular and retropharyngeal lymph nodes, and the infective BLV, in superficial cervical and mandibular lymph nodes. It was assumed that the Korean native goat was quite susceptible to BLV infection, indicating that the goat could be a good model animal for BLV.

• BMB Reports
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• v.32 no.5
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• pp.497-501
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• 1999

Biotinylation of recombinant proteins is a powerful tool for the detection and analysis of proteins of interest in a large variety of assay systems. The recent development of in vivo biotinylation techniques in E. coli has opened new possibilities for the production of site-specifically biotinylated proteins without the need for further manipulation after the isolation of the recombinantly expressed proteins. In the present study, a novel vector set was generated which allows the convenient cloning and expression of proteins of interest fused with an N-terminal in vivo biotinylated thioredoxin (TRX) protein. These vectors were derived from the previously reported pBIOTRX vector into which was incorporated part of the pBluescript II+phagemid multiple cloning site (MCS), amplified by PCR using a pair of sophisticated oligonucleotide primers. The functionality of these novel vectors was examined in this system by recombinant expression of rat transforming growth factor-$\beta$. Western-blot analysis using TRX-specific antibodies or peroxidase-conjugated streptavidin confirmed the successful induction of the fusion protein and the in vivo conjugation of biotin molecules, respectively. The convenience of molecular subcloning provided by the MCS and the effective in vivo biotinylation of proteins of interest makes this novel vector set an interesting alternative for the production of biotinylated proteins.

• Development and Reproduction
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• v.22 no.3
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• pp.283-288
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• 2018

The author carried out PCR-based genetic platform to investigate the hierarchical polar dendrogram of Euclidean genetic distances of one bastard halibut population, particularly for Paralichthys olivaceus, which was further connected with those of the other fish population, by involving with the precisely designed oligonucleotide primer sets. Eight oligonucleotides primers were used generating excessively alterating fragments, ranging in size of DNA bands from larger than approximately 100 bp to less than 2,000 bp. As regards average bandsharing value (BS) results, individuals from Hampyeong population (0.810) displayed lower bandsharing values than did individuals from Wando population (0.877). The genetic distance between individuals approved the existence of close relationship in the cluster II. Relatively, individuals of one bastard halibut population were fairly related to that of the other fish population, as shown in the polar hierarchical dendrogram of Euclidean genetic distances. The points of a noteworthy genetic distance between two P. olivaceus populations demonstrated this PCR procedure is one of the quite a few means for individuals and/or populations biological DNA investigates, for species security and proliferation of bastard halibut individuals in coastal region of the Korea.