• Korean Journal of Environmental Biology
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• v.37 no.1
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• pp.72-81
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• 2019

Bisphenol A (BPA), a representative endocrine disrupting chemicals, has adverse effects on growth, development and reproduction in aquatic organisms. The object of this study was to investigate the modulation of antioxidant enzyme-coding genes using quantitative real time RT-PCR (qRT-PCR), enzyme activity and total protein content, to understand oxidative stress responses after exposure to BPA for 48 h in brackish water flea Diaphanosoma celebensis. The BPA ($3mg\;L^{-1}$) significantly upregulated the expression of Cu/Zn-SOD, Mn-SOD, and catalase (CAT) mRNA. Three GST isoforms (GST-kappa, GST-mu, and GST-theta) mRNA levels significantly increased at the rate of $0.12mg\;L^{-1}$ of BPA. In particular, GST-mu showed the highest expression level, indicating its key role in antioxidant response to BPA. SOD activity was induced with a concentration-dependent manner, and total protein contents was reduced. These findings indicate that BPA can induce oxidative stress in this species, and these antioxidants may be involved in cellular protection against BPA exposure. This study will provide a better understanding of molecular mode of action of BPA toxicity in aquatic organisms.

• Proceedings of the Korean Society of Developmental Biology Conference
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• 2003.10a
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• pp.84-84
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• 2003

DNA methylation is involved in epigenetic processes such as X-chromosome inactivation, imprinting and silencing of transposons. DNA methylation is a highly plastic and critical component of mammalian development The DNA methyltransferases (Dnmts) are responsible for the generation of genomic methylation patterns, which lead to transcriptional silencing. The maintenance DNA methyltransferase enzyme, Dnmt 1, and the de novo methyltransferase, Dnmt3a and Dnmt3b, are indispensable for development because mice homozygous for the targeted disruption of any of these genes are not viable. The occurrence of DNA methylation is not random, and it can result in gene silencing The mechanisms underlying these processes are poorly understood. It is well established that DNA methylation and histone deacetylation operate along a common mechanistic pathway to repress transcription through the action of methyl-binding domain proteins (MBDs), which are components of, or recruit, histone deacetylase (HDAC) complexes to methylated DNA. As a basis for future studies on the role of the DNA-methyl-transferase in porcine development, we have isolated and characterized a partial cDNA coding for the porcine Dnmt1. Total RNA of testis, lung and ovary was isolated with TRlzol according to the manufacture's specifications. 5 ug of total RNA was reverse transcribed with Super Script II in the presence of porcine Dnmt 1 specific primers. Standard PCRs were performed in a total volume of 50 ul with cDNA as template. Two DNA fragmenets in different position were produced about 700bp, 1500bp and were cloned into pCR II-TOPO according to the manufacture's specification. Assembly of all sequences resulted in a cDNA from 158bp of 5'to 4861bp of 3'compare with the known human maintenance methyltransferase. Now, we are cloning the unknown Dnmt 1 region by 5'-RACE method and expression of Dnmt 1 in tissues from adult porcine animals.

• Journal of Animal Science and Technology
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• v.50 no.6
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• pp.753-762
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• 2008

The primers for RT-PCR and RACE-PCR were designed by aligning the pig genomic sequence and the human complement factor B(CFB) coding sequence(CDS) from the GenBank. Each PCR product was amplified in pig cDNA and sequencing was carried out. The CDS length of pig CFB gene was determined to be 2298 bp. In addition, the pig CDS was more longer than human and mouse orthologs because of insertion and deletion. The identities of porcine nucleotide sequences with those of human and mice were 84% and 80%, and the identities of amino acids were 79% to 77%, respectively. Three complement control protein(CCP) domains, one Von Willebrand factor A(VWFA) domain and a serine protease domain, that are revealed typically in mammals, were found in the pig CFB gene. Based on the CDSs determined, the primers were designed in intron regions for amplification of entire length of exons. In amplification and direct sequencing with genomic DNAs of six pig breeds, three cSNPs(coding single nucleotide polymorphisms) were identified and verified as missense mutations. Using the Multiplex-ARMS method, we genotyped and verified the mutations identified from direct sequencing. To demonstrate recrudescence, we performed both direct sequencing and Multiplex-ARMS with two randomly selected DNA samples. The genotype of each sample exhibited the same results using both methods. Therefore, three cSNPs were identified from pig CFB gene and that can be used for haplotype analysis of the swine leukocyte antigen(SLA) class III region. Moreover, the results indicate that the Multiplex-ARMS method should be powerful for genotyping of genes in the SLA region.

• Korean Journal of Microbiology
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• v.41 no.1
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• pp.67-73
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• 2005

Photoautotrophic bacteria are promising candidates for the production of poly(3-hydroxybutyrate) (PHB) since they can address the critical problem of substrate costs. In this study, we isolated 25 Tn5-inserted mutants of the Synechocystis sp. PCC 6803 which showed enhanced PHB accumulation compared to the wild-type strain. After 5-days cultivation under nitrogen-limited mixotrophic conditions, the intracellular levels of PHB content in these mutants reached up to $10-30\%$ of dry cell weight (DCW) comparable to $4\%$ of DCW in the wild-type strain. Using the method of inverse PCR, the affected genes of the mutants were mapped on the completely known genome sequence of Synechocystis sp. PCC 6803. As a result, the increased PHB accumulation in 5 mutants were found to be resulted from defects of genes coding for NADH-ubiquinone oxidoreductase, O-succinylbenzoic-CoA ligase, photosystem II PsbT protein or histidine kinase, which are involved in photosystem in thylakoid inner membrane of the cell. The values of $NAD(P)H/NAD(P)^+$ ratio in the cells of these mutants were much higher than that of the wild-type strain as measured by using pulse-amplitude modulated fluorometer, suggesting that PHB synthesis could be enhanced by increasing the level of cellular NAD(P)H which is a limiting substrate for NADPH-dependent acetoacetyl-CoA reductase. From these results, it is likely that NAD(P)H would be a limiting factor for PHB synthesis in Synechocystis sp. PCC 6803.

• Korean Journal of Microbiology
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• v.55 no.3
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• pp.289-292
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• 2019

Lachnospiraceae bacterium KGMB03038 (=KCTC 15821) belonging to the class Clostridia in phylum Firmicutes, was isolated from a stool sample of a healthy Korean. Herein, we report the complete genome sequence of strain KGMB03038 analyzed using the PacBio Sequel platform. The genome comprises of 3,334,474 bp with G + C content of 47.8%, which includes 3,099 predicted protein-coding genes, 12 ribosomal RNAs, 54 transfer RNAs, and 4 ncRNAs. Genome analysis revealed that strain KGMB03038 possesses a number of genes involved in hydrolysis of carbohydrates, including mono-, di-, and oligo-saccharides, and biosynthesis of various amino acids.

• Journal of Life Science
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• v.19 no.6
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• pp.765-769
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• 2009

Recent studies have reported that the distribution of HPV-16 sequence variation differs geographically, and more specifically that HPV-16 E6/E7 intratypic variants might carry a high risk for development of ICC (invasive cervical cancer) and CIN (cervical intraepithelial neoplasia) in a given population. To investigate the genetic diversities of HPV-16 E6/E7 oncogene by region, we collected nineteen HPV-16 isolates from sexually high-risk women in Busan, and analyzed the HPV-16 E6/E7 coding regions (nt 34 to 880) with HPV-16 E6/E7 specific PCR amplification. At the nucleotide levet eleven variants of the E6 genes and nine variants of the E7 genes were identified as follows: E6 T178G (n=l1), E6 T178A (n=l), E6 T350G (n=3), E6 A442C (n=2), E6 AI04T, E6 All1G, E6 C116T, E6 G145T, E6 T183G, E6 C335T, E6 G522C and E7 A647G (n=12), E7 A645C, E7 A777C, E7 G663A, E7 T732C, E7 T760C, E7 A775T, E7 T789C and E7 T795G, respectively. At the amino acid levet the isolated HPV-16 E6 and E7 genes showed eleven E6 variants: E6 D25E (n=12), E6 L83V (n=4), E6 E113D (n=2), E6 MIL, E6 Q3R, E6 P5S, E6 Q14H, E6 D25N, E6 127R, E6 H78Y, E6 C140S and three E7 variants: N29S (n=12), L28F, T72S. HPV16 E6 L83V, the dominant variant in the Caucasian population, showed relatively low frequencies in our study population. We elucidated that the dominant HPV-16 E6/E7 variants were HPV-16 E6 D25E (63.2%) and HPV-16 E7 N29S (63.2%), which were phylogenetically included in Asian lineage. Further study is needed to evaluate the risk of cervical cancer related HPV-16 E6/E7 intratypic variants in the Korean population.

• The Journal of Natural Sciences
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• v.4
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• pp.103-142
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• 1991

These studies were carried out to select somatic hybrid using selectable marker genes of Nicotiana glauca transformed by NPTII gene and Solanum tuberosum transformed by T- DNA, and to study characteristics of transformant. The results are summarized as follows. 1. Crown gall tumors and hairy roots were formed on potato tuber disc infected by A. tumefaciens Ach5 and A. rhizogenes ATCC15834. These tumors and roots could be grown on the phytohormone free media. 2. Callus formation from hairy root was prompted on the medium containing 2, 4 D 2mg/I with casein hydrolysate lg/l. 3. The survival ratio of crown gall tumor callus derived from potato increased on the medium containing the activated charcoal 0. 5-2. 0mg/I because of the preventions on the other hand, hairy roots were necrosis on the same medium. 4. Callus derived from hairy root were excellently grown for a short time by suspension culture on liquid medium containing 2, 4-D 2mg/I and casein hydrolysate lg/l. 5. The binary vector pGA643 was mobilized from E. coli MC1000 into wild type Agrobacteriurn tumefaciens Ach5, A. tumefaciens $A_4T$ and disarmed A. tuniefaciens LBA4404 using a triparental mating method with E. ccli HB1O1/pRK2013. Transconjugants were obtained on the minimal media containing tetracycline and kanamycin. pGA643 vectors were confirmed by electrophoresis on 0.7% agarose gel. 6. Kanamycin resistant calli were selected on the media supplemented with 2, 4-D 0.5mg/1 and kanamycin $100\mug$/ml after co- cultivating with tobacco stem explants and A. tumefaciens LBA4404/pGA643, and selected calli propagated on the same medium. 7. The multiple shoots were regenerated from kanamycin resistant calli on the MS medium containing BA 2mg/l. 8. Leaf segments of transformed shoot were able to grow vigorusly on the medium supplemented with high concentration of kanamycin $1000\mug$/ml. 9. Kanamycin resistant shoots were rooting and elongated on medium containing kanamycin $100\mug$/ml, but normal shoot were not. 10. For the production of protoplast from potato calli transformed by T-DNA and mesophyll tissue transformed by NPTII gene, the former was isolated in the enzyme mixture of 2.0% celluase Onozuka R-10, 1.0% dricelase, 1.0% macerozyme. and 0.5M mannitol, the latter was isolated in the enzyme mixture 1.0% Celluase Onozuka R-10, 0.3% macerozyme, and 0.7M mannitol. 11. The optimal concentrationn of mannitol in the enzyme mixture for high protoplast yield was 0.8M at both transformed tobacco mesophyll and potato callus. The viabilities of protoplast were shown above 90%, respectively. 12. Both tobacco mesophyll and potato callus protoplasts were fused by using PEG solution. Cell walls were regenerated on hormone free media supplemented with kanamycin after 5 days, and colonies were observed after 4 weeks culture.

• Journal of Plant Biotechnology
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• v.35 no.3
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• pp.177-183
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• 2008

Ac/Ds mutant lines of this study were transgenic rice plants, each of which harbored the maize transposable element Ds together with a GUS coding sequence under the control of a promoterless(Ds-GUS). We selected the mutants that were GUS expressed lines, because the GUS positive lines will be useful for identifying gene function in rice. One of these mutants was identified knock-out at Oszinc626(NP_001049991) gene, encoding a RING-H2 zinc-finger protein, by Ds insertion. In this mutant, while primary root development is normal, secondary root development from lateral root was very poor and seed development was incomplete compare with normal plant. RING zinc-finger proteins play important roles in the regulation of development in a variety of organisms. In the plant kingdom, a few genes encoding RING zinc-finger proteins have been documented with visible effects on plant growth and development. The consensus of the RING-H2(C3-H2-C3 type) domain for this group of protein is $Cys-X_2-Cys-X_{28}-Cys-X-His-X_2-His-X_2-Cys-X_{14}-Cys-X_2-Cys$. Oszinc626 encodes a predicted protein product of 445 amino acids residues with a molecular mass of 49 kDa, with a RING-zinc-finger motif located at the extreme end of the C-terminus. RT-PCR analysis indicated that the expression of Oszinc626 gene was induced by IAA, cold, dehydration, high-salinity and abscisic acid, but not by 2,4-D, and the transcription of Oszinc626 gene accumulated primarily in rice immature seeds, root meristem and shoots. The gene accumulation patterns were corresponded with GUS expression.

• The Korean Journal of Malacology
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• v.25 no.2
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• pp.153-160
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• 2009

Pisidium (Neopisidium) coreanum is a freshwater snail and lives in spring water near mountain areas. Interestingly, this snail has been traditionally regarded as medicinal food, and thus has been used as folk remedies for healing broken bones. Recently, alpha classification on Pisidium (Neopisidium) coreanum through redescription has been conducted. However, not much attention has been made in beta classification. In this study, we performed the beta classification based on metallothionein (MT) genes found from various organisms. To this end, the complete cDNA sequences were obtained from the Expressed Sequence Tag (EST) sequencing project of Pisidium (Neopisidium) coreanum. The coding region (315 bp) encoded an amino acid sequence of 105 residues. The combined results from BLAST analyses, multiple sequence alignment and molecular phylogenetic study of Pc-MT gene indicate that Pisidium (Neopisidium) coreanum has similarity to freshwater bivalve such as Dreissena polymorpha (zebra mussel), Unio tumidus (swollen river mussel) and Crassostrea ariakensis (suminoe oyster).

• Journal of The Korean Society of Inherited Metabolic disease
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• v.5 no.1
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• pp.65-75
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• 2005

Purpose: To understand genetic differences and similarities between mucolipidosis and control. Methods: Using the fibroblast of the mucolipidosis II and control, forward and reverse subtracted libraries were constructed. Among these clones, we investigated mutations in the GNPTA (MGC4170) gene, which codes for the ${\alpha}/{\beta}$ subunits of phosphotransferase, and in the GNPTAG gene, which codes for the ${\gamma}$ subunits in 5 Korean patients with mucolipidosis type II or IIIA. Result: Several differentially expressed cDNAs were cloned and their sequences were determined. Mutation analysis of the interested gene, GNPTA was performed and we identified 7 mutations in the GNPTA gene, but none in the GNPTAG gene. The mutations in type II patients included p.Q104X(c.310C>T), p.R1189X(c.3565C>T), p.S1058X(c.3173C>G), p.W894X(c.2681G>A) and p.H1158fsX15(c.3474_3475delTA), all of which are non-sense or frame shift mutations. However, a splicing site mutation, IVS13+1G>A (c.2715+1G>A) was detected along with a non-sense or a frame shift mutation (p.R1189X or p.E858fsX3(c.2574_2575delGA)) in two mucolipidosis type IIIA patients. Conclusion: This report shows that mutations in the GNPTA gene coding for the ${\alpha}{\beta}$subunits of phosphotransferase, and not mutations in the GNPTAG gene, account for most of mutations found in Korean patients with mucolipidosis type II or IIIA.