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Cloning and Expression of Bovine Polyadenylate Binding Protein 1 cDNA in Mammary Tissues

  • Kim, J.H.;Jeon, D.H.;Choi, Y.J.;Baik, M.G.
    • Asian-Australasian Journal of Animal Sciences
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    • v.14 no.6
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    • pp.771-776
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    • 2001
  • A pregnant-induced clone was identified by differential screening from a cDNA library of bovine mammary gland. The clone was identified as a cDNA encoding a polyadenylate binding protein 1 (PABP). The cDNA clone had a total length of 1,911 nucleotides coding for 636 amino acids. The nucleotide sequence of the bovine PABP was 95% and 94% identical to those of human and mouse species, respectively. Comparison of the deduced amino acid sequences of bovine PABP with those of human species showed 100% identity. Induction of the PABP mRNA was observed in bovine mammary tissues at pregnant 7 and 8 months compared to virgin, lactating and involuted states. Expression of the PABP gene was examined in mammary epithelial HC11 cells at proliferating, differentiated and apoptotic conditions. The mRNA levels of PABP gene were similar between proliferating and differentiated cells, but expression levels were very low in apoptotic cells compared to other conditions. Results demonstrate that the PABP gene is induced during pregnancy at which stage mammary epithelial cells are actively proliferating.

Mechanism of Action of Anticancer Drug Aziridinylbenzoquinones: Involvement of DT-diaphorase (DNA에 결합하는 항암제의 작용기전)

  • Lee, Chong-Soon-
    • Proceedings of the Korean Society of Applied Pharmacology
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    • 1994.11a
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    • pp.147-172
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    • 1994
  • Aziridinylbenzoquinones such as 3, 6-diaziridinyl-1, 4-benzoquinone (DZQ) and its 2, 5-methyl analog (MeDZQ) require bioreductive activation in order to elicit their anticancer activities. To determine the involvement of DTD in the activation of these drugs, we have used a ligation-mediated polymerase chain reaction to map the intracellular alkylation sites in a sing1e copy gene at the nucleotide level. We have performed this analysis in two human colon carcinoma cells, one proficient (HT-29) and one deficient (BE) in DT-diaphorase (DTD) activity. In the DTD proficient HT-29 cell line, DZQ and MeDZQ were found to alkylate both 5'-(A/T)G(C)-3' and 5'-(A/T)A-3' sequences. This is consistent with the nucleotide preferences observed when DZQ and MeDZQ are activated by purified DTD to reactive metabolites capable of alkylating DNA in vitro [Lee, C. -S., Hartley, J. A., Berardini, M. D., Butler, J., Siegel., D., Ross, D., & Gibson, N. W. (1992) Biochemistry, 31: 3019-3025]. Surprisingly in the DTD-deficient BE cell line a pattern of alkylation induced by DZQ and MeDZQ similar to that observed in the DTD-proficient HT-29 cells was observed. This suggests that reductive enzymes other than DTD can be involved in activating DZQ and MeDZQ to DNA reactive species in vivo.

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The Possibility of TBC1D21 as a Candidate Gene for Teat Numbers in Pigs

  • Jin, S.;Lee, J.B.;Kang, K.;Yoo, C.K.;Kim, B.M.;Park, H.B.;Lim, H.T.;Cho, I.C.;Maharani, D.;Lee, J.H.
    • Asian-Australasian Journal of Animal Sciences
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    • v.26 no.10
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    • pp.1374-1378
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    • 2013
  • Based on a quantitative traits locus (QTL) study using a $F_2$ intercross between Landrace and Korean native pigs, a significant QTL affecting teat numbers in SSC7 was identified. The strong positional candidate gene, TBC1D21, was selected due to its biological function for epithelial mesenchymal cell development. Sequence analysis revealed six single nucleotide polymorphisms (SNPs) in the TBC1D21 gene. Among these, two SNP markers, one silent mutation (SNP01) for g.13,050A>G and one missense mutation (SNP04) for c.829A>T (S277C), were genotyped and they showed significant associations with teat number traits (p value = 6.38E-05 for SNP01 and p value = 1.06E-07 for SNP04 with total teat numbers). Further functional validation of these SNPs could give valuable information for understanding the teat number variation in pigs.

Molecular Cloning and Expression of a Thermostable Xylose (Glucose) Isomerase Gene, xylA, from Streptomyces chibaensis J-59

  • Joo, Gil-Jae;Shin, Jae-Ho;Heo, Gun-Young;Kim, Young-Mog;Rhee, In-Koo
    • Journal of Microbiology
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    • v.43 no.1
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    • pp.34-37
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    • 2005
  • In the present study, the xylA gene encoding a thermostable xylose (glucose) isomerase was cloned from Streptomyces chibaensis J-59. The open reading frame of xylA (1167 bp) encoded a protein of 388 amino acids with a calculated molecular mass of about 43 kDa. The XylA showed high sequence homology (92% identity) with that of S. olivochromogenes. The xylose (glucose) isomerase was expressed in Escherichia coli and purified. The purified recombinant XylA had an apparent molecular mass of 45 kDa, which corresponds to the molecular mass calculated from the deduced amino acid and that of the purified wild-type enzyme. The N-terminal sequences (14 amino acid residues) of the purified protein revealed that the sequences were identical to that deduced from the DNA sequence of the xylA gene. The optimum temperature of the purified enzyme was $85^{\circ}C$ and the enzyme exhibited a high level of heat stability.

Molecular characterization and biological changes caused by Agrobacterium-mediated infiltration of PgTRX1

  • Choi, Seung Hyuk;Seo, Ji Won;Lee, Jae Geun;Yu, Chang Yeon;Seong, Eun Soo
    • Journal of Applied Biological Chemistry
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    • v.64 no.3
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    • pp.205-211
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    • 2021
  • In order to test the functionality of Panax ginseng thioredoxin 1 (PgTRX1) isolated from fermented wild ginseng roots, a transient effect on physiological activity were performed over a short time frame using the Agrobacterium infiltration technique. The PgTRX1 gene isolated from fermented wild ginseng was confirmed to have a size of 579 bp, and the expression of PgTRX1 was the highest in the sample after 6 h of fermentation. As a result of constructing this gene and confirming the infiltration reaction mediated by Agrobacterium in tobacco leaves, it was found that the expression of the NbHSR203j gene was also induced as PgTRX1 expression increased. As a result of measuring the biological activity of the infiltration samples, the total phenol content increased by 35.45±1.84 to 49.01±1.84 ㎍ GAE/mL compared to the control, and the total flavonoid amount of 9.52±0.41 to 9.82±0.25 ㎍ QE/mL was slightly high. From these results, Agrobacterium-mediated PgTRX1 appears to be related to the hypersensitive response induction mechanism of plants and the production of secondary metabolites such as phenolic substances.

Anti-metastatic mechanism of mountain cultivated wild ginseng in human cancer cell line

  • Jang, S.B.;Lim, C.S.;Jang, J.H.;Kwon, K.R.
    • Journal of Pharmacopuncture
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    • v.13 no.1
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    • pp.37-43
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    • 2010
  • Objective : Ginseng is one of most widely used herbal medicine. Ginseng showed anti-metastasis activities. However, its molecular mechanisms of action are unknown. So we want to report the wild ginseng repress which plays key roles in neoplastic epithelial-mesenchymal transition process. Methods : Treatment of the human colorectal carcinoma LOVO cells and human gastric carcinoma SNU601 cells with the increased concentrations of cultivated wild ginseng extracts resulted in a gradual decrease in the AXIN2 gene expression. Results : Metastasis-suppressor genes, maspin and nm23 was not affected by the treatment of ginseng extracts in LOVO cells. Moreover, the mountain cultivated wild ginseng or mountain wild ginseng are similar in their inhibitory effects on the expression of AXIN2 gene, but are substantially stronger than cultivated ginseng. Conclusion : We described the novel mechanism of wild ginseng-induced anti-metastasis activity by repressing the expression of AXIN2 gene that plays key roles in epithelial-mesenchymal transition process.

Detection of 881A→881G Mutation in Tyrosinase Gene and Associations with the Black Ear Coat Color in Rabbits

  • Jiang, Y.L.;Fan, X.Z.;Lu, Z.X.;Tang, H.;Xu, J.-Q.;Du, L.-X.
    • Asian-Australasian Journal of Animal Sciences
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    • v.15 no.10
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    • pp.1395-1397
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    • 2002
  • The tyrosinase gene was selected as a candidate for uncovering genetic mechanism causing 'black ear' coat color in rabbits. A PCR-SSCP detection method was established for the $881^A{\rightarrow}881^G$ mutation located in the central region of the tyrosinase gene between the CuA and CuB binding region signatures, and this was confirmed by sequencing and alignment. Fully consistent associations between the SNP and 'black ear' coat color were observed by analysis in a "black ear" pedigree and on 61 unrelated individuals. This SNP can serve as a molecular marker for use in "back ear" wool rabbit breeding.

Expression of Stat5a Gene in Bovine Mammary Gland and its Effect on Proliferation of Mammary Epithelial Cells

  • Jeon, D.H.;Choi, Y.J.;Baik, M.G.
    • Asian-Australasian Journal of Animal Sciences
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    • v.15 no.8
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    • pp.1198-1203
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    • 2002
  • To understand molecular mechanisms involved in bovine mammary gland growth, expression of stat5a gene was examined in bovine mammary tissues. We found that stat5a gene was highly induced at pregnant 7 and 8 months compared to virgin mammary tissues. To examine function of bovine stat5a in mammary epithelial cell proliferation, stat5a expression vector was transfected into mammary epithelial HC11 cells. Cell proliferation rate in stat5a gene-transfected cells was 26%, 95% and 85% higher at 24 h, 48 h and 72 h after seeding, respectively, compared to control vector-transfected cells. Results demonstrate that bovine stat5a enhances proliferation of mammary epithelial cells.

RAGE-binding peptide-conjugated polyethylenimine as a dual-functional carrier: A RAGE-mediated gene carrier and an anti-angiogenic reagent

  • Lee, Dahee;Choi, Eunji;Lee, Jaewon;Oh, Jungju;Lee, Seonyeong;Lee, Minhyung
    • Journal of Industrial and Engineering Chemistry
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    • v.67
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    • pp.284-292
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    • 2018
  • Receptor for advanced glycation end-products (RAGE) is overexpressed in various cancer cells. In this study, a RAGE-binding peptide (RBP) was conjugated to polyethylenimine (25 kDa, PEI). RBP-conjugated PEI (PEI-RBP) was characterized as a dual-functional reagent, a RAGE-mediated gene carrier and an anti-angiogenic reagent. As a gene carrier, PEI-RBP had higher transfection efficiency to the C6 glioblastoma cells than PEI. As an anti-angiogenic reagent, the pEmpty/PEI-RBP complex reduced RAGE expression on the surface of the C6 glioblastoma cells. Also, the complex reduced the VEGF expression and tube formation of endothelial cells. Therefore, PEI-RBP may be useful for development of glioblastoma therapy.

Identification of a Promoter Motif Involved in Curtovirus Sense-Gene Expression in Transgenic Arabidopsis

  • Hur, Jingyung;Choi, Eunseok;Buckley, Kenneth J.;Lee, Sukchan;Davis, Keith R.
    • Molecules and Cells
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    • v.26 no.2
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    • pp.131-139
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    • 2008
  • Expression of the seven open reading frames (ORFs) of single-stranded DNA Curtoviruses such as Beet curly top virus (BCTV) and Beet severe curly top virus (BSCTV) is driven by a bi-directional promoter. To investigate this bidirectional promoter activity with respect to viral late gene expression, transgenic Arabidopsis plants expressing a GUS reporter gene under the control of either the BCTV or BSCTV bi-directional promoter were constructed. Transgenic plants harboring constructs showed higher expression levels when the promoter of the less virulent BCTV was used than when the promoter of the more virulent BSCTV was used. In transgenic seedlings, the reporter gene constructs were expressed primarily in actively dividing tissues such as root tips and apical meristems. As the transgenic plants matured, reporter gene expression diminished but viral infection of mature transgenic plants restored reporter gene expression, particularly in transgenic plants containing BCTV virion-sense gene promoter constructs. A 30 base pair conserved late element (CLE) motif was identified that was present three times in tandem in the BCTV promoter and once in that of BSCTV. Progressive deletion of these repeats from the BCTV promoter resulted in decreased reporter gene expression, but BSCTV promoters in which one or two extra copies of this motif were inserted did not exhibit increased late gene promoter activity. These results demonstrate that Curtovirus late gene expression by virion-sense promoters depends on the developmental stage of the host plant as well as on the number of CLE motifs present in the promoter.