• Animal cells and systems
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• 제2권4호
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• pp.529-538
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• 1998

In order to analyze the intracellu1ar localization of specific RNA components of ribonucleoproteins (RNP) in Xenopus oocytes, a modified protocol of whole-mount in situ Hybridization is presented in this paper, Mitochondria specific 12S rRNA probe was used to detect the amplification and distribution of mitochondria in various stages of the oocyte life cycle, and the results were found to be consistent with previously known distribution of mitochondria. The results with other specific probes (U1 and U3 small nuclear RNAs, and 5S RNA) also indicate that this procedure is generally effective in localizing RNAs in RNP complexes even inside organelles. In addition, the RNA component of RNase MRP, the RNP with endoribo-nuclease activity, localize to the nucleus in various stages of the oocyte life cycle. Some of MRP RNA, however, were found to be localized to the special population of mitochondria near the nucleus, especially in the active stage of mitochondrial amplification. It suggests dual localization of RNase MRP in the nucleus and mitochondria, which is consistent with the proposed roles of RNase MRP in mitochondrial DNA replication and in rRNA processing in the nucleolus.

• Journal of Microbiology and Biotechnology
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• 제25권10호
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• pp.1768-1771
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• 2015

Human Fen1 protein (hFen1) plays an important role in Okazaki fragment processing by cleaving the flap structure at the junction between single-stranded (ss) DNA and doublestranded (ds) DNA, an intermediate formed during Okazaki fragment processing, resulting in ligatable nicked dsDNA. It was reported that hChlR1, a member of the cohesion establishment factor family, stimulates hFen1 nuclease activity regardless of its ATPase activity. In this study, we found that cohesion establishment factors cooperatively stimulate endonuclease activity of hFen1 in in vivo mimic condition, including replication protein-A-coated DNA and high salt. Our findings are helpful to explain how a DNA replication machinery larger than the cohesion complex goes through the cohesin ring structure on DNA during S phase in the cell cycle.

• Journal of Microbiology and Biotechnology
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• 제11권1호
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• pp.1-6
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• 2001

Ten strains of plasmid-harboring Bifidobacterium sp. were isolated from the feces of adults and children, and named as Bifidobacterium sp. GE1-GE8, ST, and SH5. These plasmids were categorized into three homologous groups (pKJ50-homologous, pKJ36-homologous, and non-homologous groups) according to Southern hybridization patterns using the formerly characterized bifidobacterial plasmids, pKJ50 and pKJ36, as probes. nine strains harboring the plasmids were shown to accumulate single-stranded DNA as a replication intermediate, based on the S1 nuclease treatment and Southern hybridization. These results suggest that the strains replicate by a rolling circle mechanism. Minimal inhibitory concentrations (MIC) of the isolated bifidobacteria against several antibiotics were determined. Two strains, GE2 and GE3, showed relatively high MiC values against tetracycline ($793.6{\mu}g/ml$) and erythromycin ($153.6{\mu}g/ml$), respectively. The tetracycline resistance of GE2 disappeared when the resident plasmid of GE2 was cured by ethidium bromide. These results show that pKJ36-homologous and pKJ50-homologous plasmids are prevalent among various Bifidobacterium strains and some Bifidobacterium plasmids appear to code for antibiotic resistance.

• Journal of Microbiology and Biotechnology
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• 제19권4호
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• pp.403-408
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• 2009

A number of bifidobacterial species of human origin were screened for the presence of cryptic plasmids. One strain, Bifidobacterium longum FI10564, harbored plasmids of approximately 2.2 kb, 3.6 kb, and 4.9 kb in size. The smallest plasmid, pFI2576(2,197 bp), was studied in detail and its complete nucleotide sequence was determined. Computer-assisted analysis of this novel plasmid(G+C content 62%) identified 9 putative open reading frames(orfs), 3 of which were shown to be probable genes. These putative genes are arranged in an operon-like structure, in which the overlapping orfs 1 and 2 encode putative Rep proteins and are highly homologous to the rep genes of the B. longum plasmid pMBI(1,847 bp). The mechanism of replication of pFI2576 was investigated using Southern blot analysis of whole cell lysates, with and without S1 nuclease treatment, and atomic force microscopy(AFM). The results indicate that pFI2576 is likely to use the theta mode of replication.

• Journal of Microbiology
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• 제34권1호
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• pp.15-22
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• 1996

The cloned X-bacterial gene (groEx) which is analogous to groE of E. Coli strongly expressed in E. coli when grown at the temperature 27.deg. C or higher without having to add any inducers. By S1-nuclease mapping, primer extension analysis and site directed mutagenesis, we found 4 promoters in the gene. Among them two promoters located at 5'-extended region of the gene are homologous to the promoters found in groE family of heat-shock genes ; they are , .sigma.$^{32}$ factor-dependent P1 promotor and .delta$^{70}$factor-dependet P2 promoter. The other two promoters found within the coding region of groESx were P3, 5'-TTGGCG-(18 bases)-AATACT-3' and P4, 5'-TTGGCA-(19 bases)-TAAGT which overlapped within 49 bases. These unique intragenic .delta.$^{70}$-dependent promoters are the first to be cloned and characterized in groE analogous heat-shock genes so far. These P3 and P4 promoters appeared to be responsible for the strong expression of GroElx in X-bacteria in vivo.

• Journal of Microbiology and Biotechnology
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• 제12권1호
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• pp.96-105
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• 2002

A bacteriocin-producing strain identified as Pediococcus acidilactici was isolated from kimchi. The bacteriocin was identified to belong to the pediocin family and exhibited bactericidal activity against most Gram-positive bacteria as well as some Gram-negative bacteria. The bacteriocin was stable up to $80^{\circ}C with wide pH ranges (5.0-10.0). The bactericidal activity remained unchanged after treatment with nonproteolytic enzymes such as nuclease and ${\alpha}$-amylase, however, it was destroyed after treatment with protease. The bacteriocin was effectively extracted by the pH-mediated adsorption-desorption method and purified effectively by semi-preparative RP-HPLC. The molecular weight of the bacteriocin was 4,622, as determined by electrospray mass spectrometry. The amino acid sequence consisted of 44 amino acid residues with four cysteines. The high solubility and pH stability of the isolated pediocin provide definite advantages over nisin and other bacteriocins in regards to its potential applications.

• 한국미생물생명공학회:학술대회논문집
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• 한국미생물생명공학회 1986년도 추계학술대회
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• pp.512.1-512
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• 1986

The cdd gene of Bacillus subtilis, encoding the deoxycytidinecytidine deaminase of pyrimidine nucleotide biosynthesis has been cloned into the EcoRl site of pBR322. The recombinant plasmid, pSol, promoted the synthesis of 100-140 fold elevated levels of the enzyme. A comparison of the polypeptides encoded by cdd complementing and non-complementing plasmids in the mini cell showed the gene product to have a molecular mass of approximately 14 kDa. The nucleotide sequence of the gene and 460 base pairs upstream from the coding region was determined. An open-reading frame, encoding a protein with a calculated molecular mass of 14337 Da, was deduced to be the coding region for cdd. However, the enzyme has an apparent molecular mass of 54 kDa as determined by gel filteration, whereas sucrose density gradient centrifugation shows 58 kDa. It means that the enzyme could be forming a tetramer in a physiological state. About 28 amino acids of the N-tetramer in a physiological state. About 28 amino acids of the N-terminal presumably form a signal for membrane translocation and six cystein residues are contained in the structure. S1 nuclease mapping indicated that transcription of cdd is initiated 17 base pairs upstream from the translational start. The structural characterization of the odd gene was performed.

• Environmental Analysis Health and Toxicology
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• 제9권1_2호
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• pp.37-51
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• 1994

To evaluate the genotoxicities of workers exposed to glue and glue cleaning solution, ambient air monitoring of working place, animal study and human monitoring were carried out. By GC-MS analysis, air samples collected from shoesmaking plant were found to be toluene, xylene, cyclohexane, n-hexane, methyl ethyl ketone, trichloroethylene, butylacetate, isopropyl alcohol. Glue and glue cleaning solution from shoesmaking plant were applicated topically to the CD-1 mice. DNA was isolated from skin 24 hr following the application and analysed for DNA-adducts using the nuclease $P_1$version of $^{32}$P-postlabelling assay. RAL (Relative Adduct Labelling, adducts$10^8$ nucleotides) was significantly increased in a dose-dependent manner in the glue cleaning solution treated mice skin. Peripheral blood DNA-adducts of workers exposed to glue and glue cleaning solution were also analysed by the same method, but there were not significant differences in the peripheral blood DNA-adducts level between exposed and control workers. In addition, glue cleaning solution from shoes factory was evaluated for mutagenicity in the Salmonella plate incorporation assay using strains TA 100 and TA 1535 in the presence and absence of Arochlor 1254-induced rat liver S$_{9}$. There was evident mutagenicity for cleaning solution in TA 100 regardless of $S_9$, but TA 1535 showed positive only in the absence of $S_9$when predicted by Stead model of mutagenicity prediction (p=0.0000). The urine concentrates from workers and controls were also assayed for mutagenicity towards strain TA 100 of Salmonella typhimurium in the presence of $S_9$ using Kado's microsuspension assay, but their mutagenic activities were not found to be significant. These data suggest that shoesmaking workers are exposed to genotoxic compounds and need to be monitored by testing the mutagenicity of human urines. However, $^{32}$P-postlabelling application requires further validation for the routine monitoring of human exposure.osure.

• 한국고분자학회:학술대회논문집
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• 한국고분자학회 2006년도 IUPAC International Symposium on Advanced Polymers for Emerging Technologies
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• pp.254-254
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• 2006

The sensitivity of plasmid DNA (pDNA) to S1 nuclease, an enzyme to cleave a single-strand DNA, was dramatically modulated through a supramolecular assembly (polyion complex micelle) with a synthetic block copolymer, poly(ethylene glycol)-b-poly(L-lysine) (PEG-PLL). The pDNA condensed in stoichiometric charge ratio was cleaved into 7 fragments each being 10/12, 9/12, 8/12, 6/12, 4/12, 3/12, and 2/12 of the original DNA length, on the other hand, the pDNA condensed in higher charge ratios (>4), were digested into non-specific manner. Condensation of the pDNA was investigated from two viewpoints that how does the rigid DNA molecules fold and condense and how does the condensation influence their biological activity.

• Laboraroty Animal Research
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• 제34권4호
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• pp.257-263
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• 2018

Trefoil factor 1 (TFF1, also known as pS2) is strongly expressed in the gastrointestinal mucosa and plays a critical role in the differentiation of gastric glands. Since approximately 50% of all human gastric cancers are associated with decreased TFF1 expression, it is considered a tumor suppressor gene. Tff1 deficiency in mice results in histological changes in the antral and pyloric gastric mucosa, with severe hyperplasia and dysplasia of epithelial cells, resulting in the development of antropyloric adenoma. Here, we generated Tff1-knockout (KO) mice, without a neomycin resistant ($Neo^R$) cassette, using the clustered regularly interspaced short palindromic repeats/CRISPR-associated nuclease 9 (CRSIPR/Cas9) system. Though our Tff1-KO mice showed phenotypes very similar to the previous embryonic stem (ES)-cell-based KO mice, they differed from the previous reports in that a reduction in body weight was observed in males. These results demonstrate that these newly established Tff1-KO mice are useful tools for investigating genetic and environmental factors influencing gastric cancer, without the effects of artificial gene insertion. Furthermore, these findings suggest a novel hypothesis that Tff1 expression influences gender differences.