• Journal of Life Science
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• v.13 no.3
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• pp.308-313
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• 2003

The collagenase gene from Vibrio parahaemolyticus 04 was subcloned into an expression vector pET-29b. The recombinant collagenase was expressed in Escherichia coli BL2l(DE3) and partially purified by Hi-Trap affinity and Sephacryl S-100 size exclusion chromatographies. The recombinant enzyme was purified by 43.7-fold and the yield was 73%. SDS-PAGE revealed that the molecular weight of the enzyme was approximately 35 kDa. Substrate specificity study of the enzyme displayed that the enzyme showed the highest activity with the type I collagen and the synthetic peptide, Z-GPGGPA, indicating that the enzyme was indeed a collagenase. The enzyme showed broad pH optimum around pH 6-12 and was stable between pH 5.5 and 11.5. The optimum temperature for the type I collagen degradation was $35^{\circ}C$. The thermostability measurement of the enzyme indicated that the enzyme was stable up to $55^{\circ}C$, but the activity was diminished quickly above $60^{\circ}C.$

• Korean Journal of Microbiology
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• v.28 no.3
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• pp.192-198
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• 1990

To determine whether the muc region of pKM101 and its mutant pSL4 is sufficient for the expression of these phenotypes, muc regions of pKM101 and pSL4 were subcloned onto the highcopy number vector pKB354 and were selected the cloned pJB200 and pJB210. The recombinant plasmids pJB200 and pJB210 were introduced into umu $C36^{-}$ uvr $A6^{-}$ (TK610) strain and determined the protection effect and mutagenecity for UV and MMS. The protection effect and mutagenecity of umu $C36^{-}$ uvr $A6^{-}$ (TK610) were supressed by muc gene of the recombinant plasmids. The muc gene of pSL4 has higher effect than that of pKM101. The recombiant plasmid pJB210(inclued muc gene of pKM101) did not affect uv-mutagenesis in the $recA^{-}$ (JC2926) mutant.

• Korean Journal of Microbiology
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• v.25 no.1
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• pp.40-45
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• 1987

The gene for the Bdi I modification enzyme, which is one of Bdi I restriction-modification system, fromBrevibacterium divaricatum FERM 5948 was cloned and expressed in E. coli. For cloning of the Bdi I methylase gene, we have initially used three cloning site(EcoRI, BamHI and Sal I) of plasmid vector pBR 322 and adopted the retransformation method after Bdi I restriction endonuclease cleavage. Selection of transformants carrying the gene was based on the resistance of the modified plasmid encoding the enzyme to cleavage by Bdi I restriction enzyme, and the recombinant plasmid pBDIM 116 containing 5.6kb EcoRI insery was proved to carry the gene. Crude cell extracts prepared from strains carrying the plasmid pBDIM 116 contained an S-adenosylmethionine-dependent methyltransferase activity specific for the Bdi I recognition site, ATCGAT. The restriction map was constructed with 11 restriction enzyme, and the Bdi I restriction-modification system was also discussed.

• KSBB Journal
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• v.10 no.3
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• pp.343-348
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• 1995

Human lipocorin-I(LCI) is a calcium ion-dependent and phospholipid-binding protein which exhibits an anti-inflammatory activity by inhibiting phospholipase A2 activity. In this study, the LCI gene containing its own terminator region was joined to GAL10 promoter-ppL (prepro-leader sequence of mating factor a). An ATG start codon of LCI gene was placed at downstream with KR endoprotease recognition site(Lys-Arg) of ppL. Recombinant S. cerevisiae harboring the LCI expression/secretion vector, pYGLPT5, was aerobicall grown on a liquid YPDG medium al $30^{\circ}C$ for 72hys. The whole cell and culture supernatant were separated after centrifugation, and the expressed LCI was analyzed by SDS-PAGE and western blotting methods. A majority fraction of the expressed LCI was found to be accumulated in the intracellular fraction, resulting in very low secretion efficiency of about 7.4%. About $500mg/\ell$ of LCI was extracellularly produced by the fed-batch culture employing the controlledfeeding of glucose and galactose. The secreted LCI was purified by ultrafiltration and hydroxylapatite column chromatography, and a purity of more than 99% was obtained.

• Microbiology and Biotechnology Letters
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• v.38 no.2
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• pp.158-162
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• 2010

The gene encoding N-terminally truncated Tod polymerase ($\Delta$Tod polymerase) from Thermus thermophilus HJ6 was expressed in Escherichia coli under the control of the lambda pR and pL tandem promoters on the expression vector pJLA503. The N-terminal domain (250 amino acids) of Tod polymerase was removed without significant effect on enzyme activity and stability, while no 5'$\rightarrow$3' exonuclease activity was detected. The $\Delta$Tod polymerase was verified to possess very efficient reverse transcriptase (RT) activity in the presence of $MgCl_2$. The cDNA can also be amplified in the polymerase chain reaction (PCR) with this mutant enzyme. The $\Delta$Tod polymerase was exhibited higher activity than the Taq polymerase in a one-step RT-PCR.

• Environmental Analysis Health and Toxicology
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• v.26
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• pp.5.1-5.11
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• 2011

Objectives: In order to identify the possibility of slender bitterling (SB) (Acheilognathus yamatsutae) being used as a test species for estrogenic endocrine disrupting chemicals (EEDCs), we carried out the cloning and sequence characterization of the estrogen receptor (ER). Methods: The ER from a slender bitterling was obtained by reverse transcriptase-polymerase chain reaction (RT-PCR), 5'- and 3'-rapid amplification of cDNA ends (5'-RACE and 3'-RACE) and T-vector cloning. The expression of ER mRNA was also analyzed in six tissues (brain, liver, kidney, gill, gonad, and intestines) by real-time PCR. Results: We obtained an ER from the slender bitterling. The SB ER cDNA was 2189 base pairs (bp) in length and contained a 1707 bp open reading frame that encoded 568 amino acid residues. The SB ER amino acid sequence clustered in a monophyletic group with the $ER{\alpha}$ of other fish, and was more closely related to zebrafish $ER{\alpha}$(88% identity) than to the $ER{\alpha}$ of other fish. The SB ER cDNA was divided into A/B, C, D, E and F domains. The SB ER has conserved important sequences for ER functions, such as the DNA binding domain (D domain), which are consistent with those of other teleosts. Conclusions: The ER of the slender bitterling could provide basic information in toxicological studies of EEDCs in the slender bitterling.

• The Journal of Korea Institute of Information, Electronics, and Communication Technology
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• v.2 no.3
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• pp.53-60
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• 2009

The vision-based driver fatigue detection is one of the most prospective commercial applications of facial expression recognition technology. The facial feature tracking is the primary technique issue in it. Current facial tracking technology faces three challenges: (1) detection failure of some or all of features due to a variety of lighting conditions and head motions; (2) multiple and non-rigid object tracking; and (3) features occlusion when the head is in oblique angles. In this paper, we propose a new active approach. First, the active IR sensor is used to robustly detect pupils under variable lighting conditions. The detected pupils are then used to predict the head motion. Furthermore, face movement is assumed to be locally smooth so that a facial feature can be tracked with a Kalman filter. The simultaneous use of the pupil constraint and the Kalman filtering greatly increases the prediction accuracy for each feature position. Feature detection is accomplished in the Gabor space with respect to the vicinity of predicted location. Local graphs consisting of identified features are extracted and used to capture the spatial relationship among detected features. Finally, a graph-based reliability propagation is proposed to tackle the occlusion problem and verify the tracking results. The experimental results show validity of our active approach to real-life facial tracking under variable lighting conditions, head orientations, and facial expressions.

• Proceedings of the Korean Institute of Information and Commucation Sciences Conference
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• 2004.05b
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• pp.603-607
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• 2004

The vision-based driver fatigue detection is one of the most prospective commercial applications of facial expression recognition technology. The facial feature tracking is the primary technique issue in it. Current facial tracking technology faces three challenges: (1) detection failure of some or all of features due to a variety of lighting conditions and head motions; (2) multiple and non-rigid object tracking and (3) features occlusion when the head is in oblique angles. In this paper, we propose a new active approach. First, the active IR sensor is used to robustly detect pupils under variable lighting conditions. The detected pupils are then used to predict the head motion. Furthermore, face movement is assumed to be locally smooth so that a facial feature can be tracked with a Kalman filter. The simultaneous use of the pupil constraint and the Kalman filtering greatly increases the prediction accuracy for each feature position. Feature detection is accomplished in the Gabor space with respect to the vicinity of predicted location. Local graphs consisting of identified features are extracted and used to capture the spatial relationship among detected features. Finally, a graph-based reliability propagation is proposed to tackle the occlusion problem and verify the tracking results. The experimental results show validity of our active approach to real-life facial tracking under variable lighting conditions, head orientations, and facial expressions.

• Korean Journal of Microbiology
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• v.40 no.1
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• pp.7-11
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• 2004

eIF5B is a translation initiation factor that delivers Met-$tRNA^{Met}$ to AUG start codon and subsequently joins the small and large ribosomes. In order to study the function of eIF5B encoded by FUN12, we constructed FUN12 which lacked 5' end of its sequence. We found that this construct lacking almost all of its promoter in pRS plasmid partially complemented slow growth phenotype of fun12 deletion strain. Interestingly, this construct expressed N-terminally truncated eIF5B and its expression level was about 5% of that of wild type eIF5B. Low amount of the eIF5B expressed additionally in fun12 deletion strain played a direct role as a limiting factor for its growth. This limiting factor eIF5B in those strains also modulates activities of overall translation in vitro.

• Korean Journal of Plant Tissue Culture
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• v.25 no.4
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• pp.225-229
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• 1998

Agrobacterium tumefaciens LBA 4404 harboring binary vector pBI 121 was used for genetic transformation of lettuce(Lactuca Sativa L.). Optimal shoot regeneration from cotyledon explants was obtained in MS medium supplemented with 0.1mg/L NAA and 1.0 mg/L 2ip. In this condition, cotyledon explants were cocultivated with A, tumefaciens for 2 days, and then transferred to selection medium supplemented with 50 mg/L kanamycin and 500 mg/L carbenicillin. These explants were subsequently subcultured every 2 weeks on shoot induction medium. PCR analysis indicated that the GUS gene was stably integrated into the nuclear genome of lettuce. Histochemical analysis based on the enzymatic activity of the CUS protein showed that GUS activity was associated with vascular tissue in leaves and roots. Progenies of Ro plants demonstrated a linked monogenic segregation for GUS gene.