• Animal cells and systems
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• v.3 no.4
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• pp.393-397
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• 1999

Length variations in human mitochondrial DNA (mtDNA) offer useful markers in the study of female aspects of human population history. One such length variation is a 9-bp deletion in the small noncoding segment located between the COII and Iysine tRNA genes (COII/tRNA/$^{Lys}$ intergenic region) which usually contain two tandemly arranged copies of a 9-bp sequence (ccccctcta) in human mtDNA. The mtDNA 9-bp deletion and polymorphic variants of expanded 9-bp repeat motif in the intergenic COII/tRNA$^{Lys}$ region have been found at varying frequencies among different human ethnic groups. We have examined the length variation of the mtDNA COII/tRNA$^{Lys}$ intergenic region from a total of 813 individuals in east Asian populations. The occurrence of the 9-bp deletion was found to be relatively homogeneous in northeast Asian populations (Chinese, 14.2%; Japanese, 14.3%: Koreans, 15.5%), with the exception of Mongolians (5.1%). In contrast, Indonesians (25.0%) and Vietnamese (23.2%) of the southeast Asian populations appeared to have relatively high frequencies of the 9-bp deletion. We identified the existence of a new expanded 9-bp repeat motif which likely resulted from a slipped mispairing insertion of six more cytosines in the intergenic COII$^{Lys}$ region. It was present at low frequencies in the Korean (2/349) and Japanese populations (2/147). Based on the results of this study, the Korean population may reflect a close genetic affinity with the Japanese and Chinese populations than the others surveyed east Asian populations.

• International Journal of Oral Biology
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• v.32 no.3
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• pp.85-91
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• 2007

Dlx3 is a homeodomain protein and is known to playa role in development and differentiation of many tissues. Deletion of four base pairs in DLX3 (NT3198) is causally related to tricho-dento-osseous (TDO) syndrome (OMIM # 190320), a genetic disorder manifested by taurodontism, hair abnormalities, and increased bone density in the cranium. Although the observed defects of TDO syndrome involves bone, little is known about the role of Dlx3 in bone remodeling process. In this study, we examined the effect of wild type DLX3 (wtDlx3) expression on osteoclast differentiation and compared it with that of 4-BP DEL DLX3 (TDO mtDlx3). To examine whether Dlx3 is expressed during RANKL-induced osteoclast differentiation, RAW264.7 cells were cultured in the presence of receptor activator of nuclear factor-B ligand (RANKL). Dlx3 protein level increased slightly after RANKL treatment for 1 day and peaked when the fusion of prefusion osteoclasts actively progressed. When wtDlx3 and TDO mtDlx3 were overexpressed in RAW264.7 cells, they enhanced RANKL-induced osteoclastogenesis and the expression of osteoclast differentiation marker genes such as calcitonin receptor, vitronectin receptor and cathepsin K. Since osteoclast differentiation is critically regulated by the balance between RANKL and osteoprotegerin (OPG), we examined the effect of Dlx3 overexpression on expression of RANKL and OPG in C2C12 cells in the presence of bone morphogenetic protein 2. Overexpression of wtDlx3 enhanced RANKL mRNA expression while slightly suppressed OPG expression. However, TDO mtDlx3 did not exert significant effects. This result suggests that inability of TDO mtDlx3 to regulate expression of RANKL and OPG may contribute to increased bone density in TDO syndrome patients. Taken together, it is suggested that Dlx3 playa role as a positive regulator of osteoclast differentiation via up-regulation of osteoclast differentiation-associated genes in osteoclasts, as well as via increasing the ratio of RANKL to OPG in osteoblastic cells.

• Korean Journal of Animal Reproduction
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• v.22 no.4
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• pp.419-424
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• 1998

The present study was carried out to examine the efficiency of cloning of transgenic embryos by nuclear transfer(NT) using gene-injected rabbit embryos. The rabbit embryos at pronuclear stage were microinjected with methallothionein-human growth hormone(MT-hGH) gene and cultured to 8- and 16-cell in TCM-199 containing 10% FCS with a monolayer of rabbit oviductal epithelial cells in a 5% $CO_2$incubator. The recipient oocytes were collected from the oviducts 14~16 h after hCG injection. The oocytes were enucleated and activated with 5$\mu$M ionomycin and 2mM 6-dimethylaminopurine. Blastomeres form gene-injected embryos were transferred into the enucleated oocytes by micromanipulation. The nuclear transplant oocytes were electrofused and co-cultured with rabbit oviductal cells. Following 120 h of culture, blastocysts were prepared for gene analysis by polymerase chain reaction(PCR). In previous experiment, the rate of gene-positive embryos detected by the nested PCR analysis was significantly decreased while developing to blastocyst(25%)(Kang et al., 1998). The fusion rate of gene-injected blastomeres was significantly(P＜0.05) lower than non-injected blastomeres(66% vs 80%). However, the NT embryos that were derived from gene-injected donor embryos did not differ from control embryos in development to the blastocyst stage(39% vs 31%). Of the 43 NT blastocysts developed from the gene-injected donor embryos, twelve(28%) were positive for the injected DNA. The results indicate that NT with gene-injected embryos can be successfully used for cloning and multiplication of transgenic embryos, furthermore applicable to improvement of transgenic animal production.

• Parasites, Hosts and Diseases
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• v.44 no.2 s.138
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• pp.117-125
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• 2006

Genetic diversity of 18 Acanthamoeba isolates from ocean sediments was evaluated by comparing mitochondrial (mt) DNA RFLP, 18S rDNA sequences and by examining their cytopathic effects on human corneal epithelial cells versus reference strains. All isolates belonged to morphologic group II. Total of 16 restriction phenotypes of mtDNA from 18 isolates demonstrated the genetic diversity of Acanthamoeba in ocean sediments. Phylogenetic analysis using 18s rDNA sequences revealed that the 18 isolates were distinct from morphological groups I and III. Fifteen isolates showed close relatedness with 17 clinical isolates and A. castellanii Castellani and formed a lineage equivalent to T4 genotype of Byers' group. Two reference strains from ocean sediment, A. hatchetti BH-2 and A. griffini S-7 clustered unequivocally with these 15 isolates. Diversity among isolates was also evident from their cytopathic effects on human corneal cells. This is the first time describing Acanthamoeba diversity in ocean sediments in Korea.

• Journal of Microbiology
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• v.37 no.2
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• pp.86-89
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• 1999

An OTHBVS cell line from HepG2 was established. This cell line stably expresses the human hepatitis B virus (HBV) middle S protein that includes the preS2 region which is important for HBV particle entry into the hepatocyte. To establish this cell line, the middle S open reading frame (ORF), with a promoter located in the 5' region and enhancer located in the 3' region, was cloned downstream from the metallothionine (MT) promoter of the OT1529 vector. In this vector, expression of the middle S protein was constructed to be regulated by its own promoter and enhancer. Expression of the large S protein which contains the preS1 region in addition to the middle S protein was designed to be regulated by the MT promoter. When extracts of OTHBVS cells were examined with an S protein detection kit (RPHA, Korea Green Cross Co.), an S protein was detected. Total mRNA of OTHBVS cell examined by northern blot analysis with an S ORF probe revealed small/middle S transcripts (2.1 kb). When the MT promoter was induced by Zn, large S transcripts (2.4 kb) were detected. The GP36 and GP33 middle S proteins were presumably detected, but large S proteins were not detected by immunostain analysis using anti-preS2 antibody.

• Animal cells and systems
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• v.4 no.1
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• pp.31-37
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• 2000

The genetic variations among six species of Rana from Korea (R. nigro-maculata, R. piancyi, R. dybowskii, R. sp, R. rugosa type A, B and R. amurensis) were investigated using 499 bases of mitochondrial DNA sequences for ND2 (NADH dehydrogenase subunit 2) gene and $tRNA^{Trp}$ gene. Partial sequences of ND2 gene (427 bp) and full sequences of $tRNA^{Trp}$ gene (73 bp) were identified. The level of sequence divergences ranged from 0.2 to 5.2% within species and 4.9-28.0% among 6 species of the genus Rana. The $tRNA^{Trp}$ gene of the genus Rana was composed of 77 nucleotides which showed a two dimensional "cloverleaf" structure. The secondary structure of $tRNA^{Trp}$ was not found compensatory changes which could potentially confound phylogenetic inference. In the neighborjoining tree, brown frogs were clustered first with the level of sequence divergence of 13.20% between R. amurensis and R. dybowskii, and 9% between R. dybowskii and R. sp. supported by 99% bootstrap iterations, respectively. R. nigromaculata and R. plancyi were clustered into another group with 5.1% divergence supported by 100% bootstrap iteration. R. rugosa A 8nd B types were grouped by 4.9% divergence and clustered into the last group with other two groups with 100% bootstrap iterations.

• The Korean Journal of Blood Transfusion
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• v.29 no.3
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• pp.253-261
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• 2018

Background: A massive blood transfusion (MT) requires significant efforts by the Blood Bank. This study examined blood product use in MT and emergency O Rh Positive red cells (O RBCs) available directly for emergency patients from the Trauma Center in Ajou University Hospital. Methods: MT was defined as a transfusion of 10 or more RBCs within 24 hours. The extracted data for the total RBCs, fresh frozen plasma (FFP), platelets (PLTs, single donor platelets (SDP) and random platelet concentrates (PC)) issued from Blood Bank between March 2016 and November 2017 from Hospital Information System were reviewed. SDP was considered equivalent to 6 units of PC. Results: A total of 345 MTs, and 6233/53268 (11.7%) RBCs, 4717/19376 (24.3%) FFP, and 4473/94166 (4.8%) PLTs were used in MT (P<0.001). For the RBC products in MT and non-MT transfusions, 28.0% and 34.1% were group A; 27.1% and 26.0% were group B; 37.3% and 29.7% were group O, and 7.5% and 10.2% were group AB (P<0.001). The ratios of RBC:FFP:PLT use were 1:0.76:0.72 in MT and 1:0.31:1.91 in non-MT (P<0.001). A total of 461 O RBCs were used in 36.2% (125/345) of MT cases and the number of O RBCs transfused per patient ranged from 1 to 18. Conclusion: RBCs with the O blood group are most used for MT. Ongoing education of clinicians to minimize the overuse of emergency O RBCs in MT is required. A procedure to have thawed plasma readily available in MT appears to be of importance because FFP was used frequently in MT.

• Microbiology and Biotechnology Letters
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• v.17 no.5
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• pp.454-460
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• 1989

Antibiotics resistance genes both in natural bacterial isolates and the genetically cloned bacteria were comparatively studied for their transfer frequencies by the method of conjugation in several different water environments. The Kmr genes in both kinds of bacteria were transferred more frequently in autoclaved wastewater of laboratory environment than in natural river water, but in Luria Bertani (LB) broth medium under the laboratory conditions the transfer frequences of the genes were much higher than in the autoclaved wastewater. The transfer frequencies at 2$0^{\circ}C$ and 3$0^{\circ}C$ were not much different in any water environments. The Km$^{${\gamma}$}$ genes of the genetically cloned bacteria and the natural isolates were transferred at the same frequency both in natural river water and in the autoclaved wastewater of laboratory environment, but in LB broth under laboratory conditions the transfer frequencies were lowered by 10$^{-3}$ to 10$^{-4}$ in the genetically cloned cells than the natural isolates. When donors of different cloned cells were conjugated with recipient of a natural isolates, the Km$^{${\gamma}$}$ genes of different donor cells were transferred at the about same frequency, but the same donor of the cloned cell were conjugated with recipients of different natural isolates, the transfer of Km$^{${\gamma}$}$ gene of the cloned cell showed some differences of 101 to 102 in frequency.

• Journal of Animal Reproduction and Biotechnology
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• v.34 no.1
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• pp.10-19
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• 2019

Morphology of cumulus-oocyte-complexes (COCs) at germinal vesicle (GV) stage as one of the evaluation criteria for oocyte maturation quality after in vitro maturation (IVM) plays important roles on the meiotic maturation, fertilization and early embryonic development in pigs. When cumulus cells of COCs are insufficient, which is induced the low oocyte maturation rate by the increasing of reactive oxygen species (ROS) in porcine oocyte during IVM. The ROS are known to generate including superoxide and hydrogen peroxide from electron transport system of mitochondria during oocyte maturation in pigs. To regulate the ROS production, the cumulus cells is secreted the various antioxidant enzymes during IVM of porcine oocyte. Our previous study showed that Mito-TEMPO, superoxide specific scavenger, improves the embryonic developmental competence and blastocyst formation rate by regulating of mitochondria functions in pigs. However, the effects of Mito-TEMPO as a superoxide scavenger to help the anti-oxidant functions from cumulus cells of COCs on meiotic maturation during porcine oocyte IVM has not been reported. Here, we categorized experimental groups into two groups (Grade 1: G1; high cumulus cells and Grade 2: G2; low cumulus cells) by using hemocytometer. The meiotic maturation rate from G2 was significantly (p < 0.05) decreased (G1: $79.9{\pm}3.8%$ vs G2: $57.5{\pm}4.6%$) compared to G1. To investigate the production of mitochondria derived superoxide, we used the mitochondrial superoxide dye, Mito-SOX. Red fluorescence of Mito-SOX detected superoxide was significantly (p < 0.05) increased in COCs of G2 compared with G1. And, we examined expression levels of genes associated with mitochondrial antioxidant such as SOD1, SOD2 and PRDX3 using a RT-PCR in porcine COCs at 44 h of IVM. The mRNA levels of three antioxidant enzymes expression in COCs from G2 were significantly (p < 0.05) lower than COCs of G1. In addition, we investigated the anti-oxidative effects of Mito-TEMPO on meiotic maturation of porcine oocyte from G1 and G2. Meiotic maturation and mRNA levels of antioxidant enzymes were significantly (p < 0.05) recovered in G2 by Mito-TEMPO ($0.1{\mu}M$, MT) treatment (G2: $68.4{\pm}3.2%$ vs G2 + MT: $73.9{\pm}1.4%$). Therefore, our results suggest that reduction of mitochondria derived superoxide by Mito-TEMPO may improves the meiotic maturation in IVM of porcine oocyte.

• Animal cells and systems
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• v.4 no.3
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• pp.267-272
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• 2000

Partial sequence of the mitochondrial cytochrome b gene was analyzed to investigate genetic variation from 10 geographic populations of Granulilittorina exigua in Korea. The sequence of 282 base pairs was determined by PCR-directed silver sequencing method. The sequences of two species within the genus Littorina reserved in NIH blast search were utilized to determine geographic variations of species referred. The levels of mtDNA sequence differences were 0.00-2.54% within populations and 0.71-4.43% between populations. There were four amino acid differences between representative species of the genera Granulilittorina and Littorina, but no differences within populations of the genus Granulilittorina. The UPGMA and the N-J trees based on Tamura-Nei genetic distance matrix were constructed, which showed that the genus Granulilittorina was divided into three groups such as eastern (even exception for Tokdo population), southern, and western regional populations. The degrees of genetic divergence within populations of each group were p=0.021, p=0.019, and p=0.018, respectively. The divergence between the eastern and southern populations was p=0.032, showing closer relationship than with the western populations (p=0.052). Based on the diverged time estimation, the eastern and southern populations of Granulilittorina exigua in Korea diverged from the western populations about 2.1 MYBP, and the eastern and southern populations diverged from each other about 1.3 MYBP.