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Molecular Cloning and Expression of Human Poly (ADP-ribose) Synthetase cDNA in E. Coli (인간 Poly(ADP-ribose) Synthetase cDNA의 클로닝 및 대장균에서의 발현)

  • 이성용;김완주;이태성;박상대;이정섭;박종군
    • The Korean Journal of Zoology
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    • v.39 no.3
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    • pp.248-256
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    • 1996
  • The present study was performed to clone and express human poly (ADP-ribose) synthetase (PARS) cDNA in E coli. For these purposes, the CDNA for human poly (ADP-ribose) synthetase, encoding the entire protein, was cloned into pGEM-7Zf(+). The resulting recombinant plasmid pPARS6.1 was restriction enzyme mapped and its identity was confirmed by Southern blot analysis. The pPARS6. 1 contained full-length CDNA of human PARS and the nudeotide sequences were identical with those reported previously. The recombinant protein which migrated as a unique 120 kDa band on 10% SDS-polyacrylamide gels, was identified as PARS by Southwestern blots using nick-translated DNA probes and by activity gels and activity blots using 32 P-NAD as a substrate for poly (ADP-ribose) synthetase (PARS). The signals corresponding to 120 and 98 kDa proteins were obtained following IPTG (0.4 mM) induction of the PARS cDNA cloned into Xba I-digested pGEM-7Zf(+) vector. Nonspecific signals corresponding to 45 and 38 kDa proteins were also shown in both IPTG-induced and noninduced cells. The nonspecific proteins may be products of incomplete translation or proteolytic products of intact PARS.

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Identification of Sex-specific Expression Markers in the Giant Tiger Shrimp (Penaeus monodon)

  • Khamnamtong, Bavornlak;Thumrungtanakit, Supaporn;Klinbunga, Sirawut;Aoki, Takashi;Hirono, Ikuo;Menasveta, Piamsak
    • BMB Reports
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    • v.39 no.1
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    • pp.37-45
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    • 2006
  • Bulked segregant analysis (BSA) and AFLP were used for isolation of genomic sex determination markers in Penaeus monodon. A total of 256 primer combinations were tested against 6-10 bulked genomic DNA of P. monodon. Five and one candidate female- and male-specific AFLP fragments were identified. Female-specific fragments were cloned and further characterized. SCAR markers derived from FE10M9520, FE10M10725.1, FE10M10725.2 and FE14M16340 provided the positive amplification product in both male and female P. monodon. Further analysis of these markers using SSCP and genome walk analysis indicated that they were not sex-linked. In addition, sex-specific (or differential) expression markers in ovaries and testes of P. monodon were analyzed by RAP-PCR (150 primer combinations). Twenty-one and fourteen RAP-PCR fragments specifically/differentially expressed in ovaries and testes of P. monodon were successfully cloned and sequenced. Expression patterns of 25 transcripts were tested against the first stranded cDNA of ovaries and testes of 3-month-old and broodstock-sized P. monodon (N = 5 and N = 7 - 10 for females and N = 4 and N = 5 - 7 for males, respectively). Five (FI-4, FI-44, FIII-4, FIII-39 and FIII-58) and two (M457-A01 and MII-51) derived RAP-PCR markers revealed female- and male-specific expression patterns in P. monodon. Surprisingly, MII-5 originally found in testes showed a higher expression level in ovaries than did testes of juvenile shrimps but a temporal female-specific pattern in P. monodon adults.

Comparison of the [$^2H_5$]Phenylalanine Model with the [1-$^{13}C$]Leucine Method to Determine Whole Body Protein Synthesis and Degradation in Sheep Fed at Two Levels

  • Al-Mamun, M.;Ito, C.;Fujita, T.;Sano, H.;Sato, A.
    • Asian-Australasian Journal of Animal Sciences
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    • v.20 no.10
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    • pp.1517-1524
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    • 2007
  • The [$^2H_5$]phenylalanine model was compared with the [1-$^{13}C$]leucine method to determine whole body protein synthesis (WBPS) and degradation (WBPD) in sheep fed at two levels. The animals were fed either 103 (M-diet) or 151 (H-diet) kcal $ME/kg^{0.75}/day$ once daily in a crossover design for 21 days each. The isotope dilutions were simultaneously conducted as a primed-continuous infusion of [$^2H_5$]phenylalanine, [$^2H_2$]tyrosine and [1-$^{13}C$]leucine on each dietary treatment. The WBPS and WBPD calculated from the [$^2H_5$]phenylalanine model were lower (p = 0.009 and p = 0.003, respectively) than those calculated from the [1-$^{13}C$]leucine method. The WBPS tended to be higher (p = 0.08) and WBPD was numerically higher (p = 0.33) for H-diet than M-diet in the [$^2H_5$]phenylalanine model, whereas the WBPS was numerically higher (p = 0.37) for H-diet and WBPS remained similar (p = 0.79) between diets in the [1-$^{13}C$]leucine method. However, the absolute values and the directions of WBPS as well as WBPD from M-diet to H-diet were comparable between the [$^2H_5$]phenylalanine model and [1-$^{13}C$]leucine method. Moreover, the values vary depending on the use of the respective amino acid contents in the carcass protein when calculating WBPS and WBPD. Therefore, it is concluded that the [$^2H_5$]phenylalanine model could be used as an alternative to the [1-$^{13}C$]leucine method for the determination of WBPS and WBPD in sheep.

The Role of Nostalgia in Self-congruence Consumption (자아일치 소비에 대한 노스탤지어의 역할)

  • Kim, Dong Kyu;Chun, Seungwoo;Park, Joon Woo;Hwang, Chunduk
    • The Journal of the Korea Contents Association
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    • v.16 no.10
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    • pp.373-381
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    • 2016
  • This study examined the interaction of nostalgia and self-congruence on consumer behavior. We proposed that nostalgia make consumers' brand attitude and purchase intention more positive when the level of congruence between the brand and consumer's ideal self-image is high, irregardless of the level of actual self congruence. 283 middle-aged people participated and 2(nostalgia: stimuli vs. control) X 2(ideal self-congruence: high vs. low) between subject design is used. The result showed the interaction effect of nostalgia and ideal self congruence. Nostalgia-primed (vs. control) participants reported higher brand attitude and purchase intention to the ideal self-congruent brand. Finally the theoretcal and practical implications and limitations of this study are discussed.

Stage-specific Expression of Lanosterol 14${\alpha}$-Demethylase in Mouse Oocytes in Relation to Fertilization and Embryo Development Competence

  • Song, Xiaoming;Ouyang, Hong;Tai, Ping;Chen, Xiufen;Xu, Baoshan;Yan, Jun;Xia, Guoliang;Zhang, Meijia
    • Asian-Australasian Journal of Animal Sciences
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    • v.22 no.3
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    • pp.319-327
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    • 2009
  • Follicular fluid meiosis-activating sterol (FF-MAS) has been suggested as a positive factor which could improve the oocyte quality and subsequent embryo development after in vitro fertilization. However, FF-MAS is a highly lipophilic substance and is hard to detect in studying the relationship between MAS and quality of oocyte maturation. The present study focused on the expression of lanosterol 14${\alpha}$-demethylase (LDM), a key enzyme that converts lanosterol to FF-MAS, on mouse oocyte maturation and its potency on development. LDM expression was strong in gonadotropin-primed germinal vesicle stage oocytes, weak after germinal vesicle breakdown (GVBD), and then strong in MII stage oocytes. The LDM-specific inhibitor azalanstat significantly inhibited oocyte fertilization (from 79.4% to 68.3%, p<0.05). Also, azalanstat (5 to 50 ${\mu}M$) decreased the percentage of blastocyst development dosedependently (from 78.7% to 23.4%, p<0.05). The specific inhibition of sterol ${\Delta}14$-reductase and ${\Delta}7$-reductase by AY9944 accumulates FF-MAS and could increase blastocyst development rates. Additionally, in the AY9944 group, the rate of inner cell mass (ICM)/ total cells was similar to that of in vivo development, but the rate was significantly decreased in azalanstat treatment. In conclusion, LDM, the key enzyme of FF-MAS production, may play an important role in fertilization and early development of the mouse embryo, especially in vitro.

The Cerebral Activation of the Emotional and Linguistic Attributes during Visual Word Recognition: fMRI Study (시각 단어 재인동안 정서적 속성과 언어적 속성에 의해 활성화되는 대뇌 영역 : fMRI 연구)

  • Park, Chang-Su;Han, Jong-Hye;Choi, Moon-Gee;Nam, Ki-Chun
    • Proceedings of the Korean Society for Cognitive Science Conference
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    • 2006.06a
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    • pp.53-58
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    • 2006
  • We examined the cerebral activation of the emotional and linguistic attributes during the visual word recognition. This research investigated the affective priming effect preserving the behavioral paradigm. We used the primed-evaluation task in which the participants classify the target as positive or negative, and manipulated the emtional attributes by emtional relations of the prime-target word pairs(PP, PN, NP, NN). ROIs analyses for the semantic processing and emotional processing were performed. The results showed that the semantic processing areas including the IPL, SMG, and aSTS were activated differently according to the experimental condition. The activations of the IPL were increased only on the NN condition, whereas the activation of the SMG was decreased only on the PP condition. Furthmore, the activation of the emotional processing areas including the mPFC and ACC, was different according to the emotional realtions of word pairs. Similar to the SMG, the BOLD signal of the mPFC was decreaed only on the PP condition, whereas the activation of ACC was Increased only on the NN condition. These results were seemed to show the interact ive cerebral activations for processing the emtoional and linguistic attributes in a word, during visual word recognition.

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Generation of FISH Probes Using Laser Microbeam Microdissection and Application to Clinical Molecular Cytogenetics

  • Shim, Sung-Han;Kyhm, Jee-Hong;Chung, Sung-Ro;Kim, Seung-Ryong;Park, Moon-Il;Lee, Chul-Hoon;Cho, Youl-Hee
    • Journal of Microbiology and Biotechnology
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    • v.17 no.7
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    • pp.1079-1082
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    • 2007
  • Chromosome microdissection and the reverse FISH technique is one of the most useful methods for the identification of structurally abnormal chromosomes. In particular, the laser microbeam microdissection (LMM) method allows rapid isolation of a target chromosome or a specific region of chromosomes without damage of genetic materials and contamination. Isolated chromosomes were directly amplified by the degenerate oligonucleotide-primed polymerase chain reaction (DOP-PCR), and then the FISH probes labeled with spectrum green- or spectrum red-dUTP were generated by nick-translation. Whole chromosome painting (WCP) probes were successfully generated from only 5 copies of the chromosome. With this method, we produced 24 WCP probes for each human chromosome. We also tried to characterize a marker chromosome, which seemed to be originated from chromosome 11 on conventional banding technique. The marker chromosomes were isolated by the LMM method and analyzed by reverse FISH. We elucidated that the marker chromosome was originated from the short arm of chromosome 5 ($5p11{\to}pter$). A fully automated and computer-controlled LMM method is a very simple laboratory procedure, and enables rapid and precise characterization of various chromosome abnormalities.

The Difference of Emotional Evaluation for Personal Pronoun 'I' and 'You' (인칭 대명사 '나'와 '너'의 정서적 평가 차이)

  • Lee, Jae-Ho
    • Korean Journal of Cognitive Science
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    • v.23 no.3
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    • pp.323-348
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    • 2012
  • Three experiments were conducted to explore the interaction of personal pronoun (e.g. 'I' and 'you') and emotional evaluation (e.g. positive and negative) using time-course (e.g. SOA 500-1000ms) and multi-task approaches (e.g. lexical decision task and primed naming task). In Experiment 1, Participants were presented personal pronoun as primes at SOA 1000ms and were asked to response emotional words which were differed in emotional attributes. The results showed that the interaction effects of personal pronoun and emotional words were found. In Experiment 2, Participants were presented personal pronoun as primes at SOA 1000ms and were asked to response emotional words which were differed in emotional attributes. The results showed that no effects were found. In Experiment 3, Participants were presented personal pronoun as primes at SOA 500ms and were asked to pronounce emotional words which were differed in emotional attributes. The results showed that the interaction of personal pronoun and emotional words were found. The results of 3 experiments were discussed from a point of view of dynamic processes of social cognition.

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The protective efficacy of the enterotoxigenic Escherichia coli vaccine candidate by GI24 against neonatal piglet colibacillosis

  • Choi, Yeong Hwan;Moon, Ja Young;Seo, Byoung Joo;Kim, Won Kyung;Cho, Jeong Sang;Choi, Min Su;Lim, Jae Sam;Kim, Sung Bok;Kim, Won Il;Hur, Jin
    • Korean Journal of Veterinary Service
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    • v.40 no.4
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    • pp.235-244
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    • 2017
  • Enterotoxigenic Escherichia coli (ETEC) strains producing each F4, F5, F6 and F41 fimbriae were lysed by GI24 peptide. The lysate cells were used as ETEC vaccine candidate. This study was carried out to examine whether intramuscular (im) immunization of pregnant sows with the novel vaccine candidate could effectively protect their neonatal piglets against ETEC colibacillosis. All pregnant sows were primed at 11 weeks and were boosted at 14 weeks of pregnancy. Group A sows were im inoculated with PBS. Group B sows were im immunized with $2{\times}10^9$ the mixture. Seral IgG, colostral IgA and IgG titers from group B sows, and seral IgG and IgA levels in group B piglets were significantly higher than those of group A sows and piglets, respectively. After challenge with wild-type ETEC, diarrhea and mortality was not observed in group B piglets. However, diarrhea was observed in 66.7% of group A piglets, and 33.3% mortality was observed. These findings indicate that im immunization of sows with the mixture of the novel vaccine candidate can effectively protect their offspring from ETEC colibacillosis.

Effects of Osmotic and Non-Osmotic Priming Methods for Enhancing Germination of Pepper Seed

  • Kim, Min Geun;Kang, Won Sik;Kim, Du Hyun
    • Proceedings of the Plant Resources Society of Korea Conference
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    • 2018.10a
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    • pp.87-87
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    • 2018
  • This study was conducted to investigate the effects of osmotic priming, liquid smoke (LS), sonication and modified drum priming treatments to improve the germination speed and uniformity of pepper seeds for high quality export seeds. Seeds were treated in 0.5, 1.0, 5.0, 10% LS solution only or with 100mM $K_2SO_4$ solution for 6days at $15^{\circ}C$. Sonication treatment was performed for 5, 10, and 20 minutes at an intensity of 5.2, 10.4, 15.7, 21.0, 26.1kHz in water at $15^{\circ}C$. After sonication treatment, seeds were primed with water or 100mM $K_2SO_4$ for 4days. 40, 50 and 60% seed moisture content (SMC) of hydrated seeds were incubated for 84, 96 and 108h in a container with a relative humidity of 99% at 26rpm for a modified drum priming treatments. 0.5% LS treatment with water or 100mM $K_2SO_4$ priming showed significant effects with 71% GP (9.0days MGT) and 66% GP (7.4days MGT), respective, while untreated seeds resulted in only 61% GP and 10days MGT. The healthy seed % was increased by 24% by $K_2SO_4$ priming with LS treatment compared to untreated seeds. However, sonication treatment showed less germination or no difference compared to untreated seed, regardless of intensity and treatment time. The modified drum priming treatment (108h incubation after 50% SMC hydration) significantly improved the GP (on the $4^{th}$days after sowing), germination rate (GR), and MGT to 65%, $28%{\cdot}day^{-1}$, and 3.6days, whereas untreated seed resulted only 7% GP, $17%{\cdot}day^{-1}$ GR and 5.9days MGT. Thus, modified drum priming treatment effectively improved seed germination and is considered as an industrially promising treatment methods considering the shortening of the treatment period and environment-friendly aspects.

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