• 미생물학회지
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• 제30권1호
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• pp.8-14
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• 1992

Pseudomonas sp. DJ77로부터 extradiol dioxygenase 유전자(phnE)를 클로닝하고 염기배열을 결정하였다. 921 bp의 open reading frame (ORF) 이 존재하였고 개시코돈 앞에서 Shine-Dalgarno sequence를 발견하였다. phnE 유전자에서 만들어지는 PhnE 단백질은 분자량이 34,449 Da 인데 SDS-polycrylamide gel 전기영동에 의해 측정된 분자량과 일치하였다. PhnE는 NahH, XylE, DmpB 등과 아미노산 배열의 상동성의 약 50% 였다. DJ77에는 bphC와 같은 3형의 extradiol dioxygenase 유전자는 발견할 수 없었다. DJ77 과 JM101(pPE17)은 catechol, 3-methylcatechol, 4-methylcatechol, 2, 3-dihydroxybiphenyl 등의 기질을 meta-cleavage 하여 노란색 화합물을 생성할 수 있었다.

• 대한생식의학회:학술대회논문집
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• 대한불임학회 2006년도 The 5th Biannual Meeting of Pacific Rim Society for Fertility and Sterility
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• pp.155.2-155.2
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• 2006

• 한국자기공명학회논문지
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• 제27권2호
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• pp.5-9
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• 2023

t ENOD40B, a plant peptide hormone, was doubly labeled with C-13 and N-15 by recombinant production in Escherichia coli. The peptide was prepared by affinity chromatography followed by protease cleavage and reverse-phase chromatography. To elucidate the mode of action against its receptor, sucrose synthase, we proceeded to assign the backbone and side-chain resonances using a set of double and triple resonance experiments. This result will be used to determine the three-dimensional structure of the peptide at its bound state as well as to observe the chemical shift changes upon binding.

• 한국동물번식학회:학술대회논문집
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• 한국동물번식학회 2001년도 춘계학술발표대회
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• pp.60-60
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• 2001

Human thrombopoietin (hTPO) is a cytokine that plays a central role in megakaryopoiesis by influencing on the development and maturation of megakaryocyte and platelet production. To induce hTPO production in the mammary gland, expression vector was constructed by combining the promoter of bovine beta-casein gene, cDNA of hTPO and neomycine resistance gene for transfection into fibroblasts. Bovine fibroblast cells derived from female ear skin were transfercted with the expression vector using Lipofectamine (Life Technology, NY). Transected cells resistant to G4l8 treatment (600 $\mu\textrm{g}$/$m\ell$) were recovered and colony formation was initiated at 13 days. The colonies with about 1 cm diameter were picked and analysed by PCR. Single transfected cells were individually transferred to enucleated oocytes. After electrofusion, the reconstructed embryos were exposed to calcium ionophore (5uM) for 5 min followed by treatment with 6-DMAP (2.5 mM) for 4h. The nuclear transfer embryos were cultured in CRlaa medium at 38.5C, 5% $CO_2$ for 7 days. Twenty three of 29 (79.3%) colonies were proved to be hTPO transfectants by PCR. The colonies were further passaged and used to produce transgenic embryos using nuclear transfer. Cleavage and developmental rates of reconstructed embryos to the blastocyst stage were 65.1% and 39.4%, respectively Of 22 blastocysts that developed from reconstructed embryos with the transfected cell, 20 embryos (90.9%) were positive for hTPO by using PCR analysis. The results suggest that somatic cell nuclear transfer is efficient for production of transgenic embryos.

• 한국가축번식학회지
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• 제20권4호
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• pp.459-465
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• 1997

본 연구는 핵이식의 시기가 융합과 체외발달에 미치는 영향을 조사하고 체외수정 공핵배의 핵과 체외성숙난자간에 전기융합의 최적 통전전압과 통전시간을 결정하기 위하여 시도하였다. 수핵난자는 체외성숙후 25~27시간에 제핵하였고 난자의 추가성숙을 위하여 융합전에 18~20시간 더 배양하였다. 공핵체외수정란은 16~32세포기까지 BOEC와 공배양하였다. 공핵배의 할구 이식시간은 조기핵이식구에서는 체핵후 1~3시간, 자연핵이식구를 체핵후 16~18시간이었다. 융합은 체외성숙후 43~45시간에 행하였다. 융합율은 할구 이식시기에 따라 차이가 없었으나 난할율과 8~16세포기 발달은 자연핵이식구에서 높았다. 융합, 난할 및 상실배 배반포 발달율은 1.0kV/cm보다 0.75kV/cm전압에서 높았으며 0.7kV/cm에서 상실배 배반포율은 17.6%였다. 통전시간에 따라 융합율은 차이가 없었으나 50$\mu$sec와 70$\mu$sec에서 난할율과 상실배-배반포 발달율이 다소 높았다. 본 결과에서 핵이식의 최적시기는 수핵난자의 체외성숙후 42~44시간이었으며 최적 전기융합 조건은 50~70$\mu$sec의 1회 0.75kV/cm임을 알 수 있었다.

• 멤브레인
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• 제13권2호
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• pp.110-117
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• 2003

술폰화 폴리이미드는 클로로알칼리 전기분해, 양이온교환수지 및 연료전지용 고분자진해질막 등과 같은 많은 응용에 있어 유용한 재료로서 연구되어지고 있다. 그러나, 이러한 응용이 이루어지는 과정에 있어서 시간에 따라 연속적인 탈수공정이나 고분자의 분해에 따른 성능 감소 등이 보고되었다. 술폰화 고분자 분해의 주요 원인 중 하나로서 고분자 분자량의 감소 및 고분자 사슬의 절단으로 이끌어지는 가수분해를 들 수 있다. 따라서, 본 연구의 목적은 수화조건 하에서의 $-SO_3$H와 연결된 이미드 사이클과 부가적인 에스테르 결합의 분해를 조사하는 데 있다. 사슬의 분리에 대해 가능한 정확한 정보를 얻고 이를 확인하기 위해서는 $^1H$ 및 $^{13}C$ NMR, FT-IR 분석을 이용했으며, 또한 보다 편리한 분석을 위해서 model compound를 사용하여 실험을 수행하였다. 결과적으로, 술폰화 폴리이미드의 수화안정성을 평가하기 위해서 프탈계 및 나프탈렌계 이미드 고리와 에스테르 결합을 갖는 model compound를 합성하였고, 제조된 model compound를 이용하여 $80^{\circ}C$ 초순수 하에서 aging 실험을 수행하였고, lyophilization technique을 사용하여 반응을 중지시켰다. Aging된 product는 NMR, FT-IR spectroscopy를 이용하여 분석하였다.

• Bulletin of the Korean Chemical Society
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• 제33권10호
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• pp.3248-3252
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• 2012

Vibrio extracellular metalloprotease (vEP), secreted from Vibrio vulnificus, shows various proteolytic function such as prothrombin activation and fibrinolytic activities. Premature form of vEP has an N-terminal (nPP) and a C-terminal (C-ter100) region. The nPP and C-ter100 regions are autocleaved for the matured metalloprotease activity. It has been proposed that two regions play a key role in regulating enzymatic activity of vEP. Especially, C-ter100 has a regulatory function on proteolytic activity of vEP. C-ter100 domain has been cloned into the E. coli expression vectors, pET32a and pGEX 4T-1 with TEV protease cleavage site and purified using gel-filtration chromatography followed by affinity chromatography. To understand how C-ter100 modulates proteolytic activity of vEP, structural studies were performed by heteronuclar multi-dimensional NMR spectroscopy. Backbone $^1H$, $^{15}N$ and $^{13}C$ resonances were assigned by data from standard triple resonance and HCCH-TOCSY experiments. The secondary structures of vEP C-ter100 were determined by TALOS+ and CSI software based on hydrogen/deuterium exchange. NMR data show that C-ter100 of vEP forms a ${\beta}$-barrel structure consisting of eight ${\beta}$-strands.

• Mass Spectrometry Letters
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• 제3권2호
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• pp.47-49
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• 2012

Myoglobin was hydrolyzed by microwave-assisted weak acid hydrolysis with 2% formic acid at $37^{\circ}C$, $50^{\circ}C$, and $100^{\circ}C$ for 1 h. The most effective hydrolysis was observed at $100^{\circ}C$. Hydrolysis products were investigated using matrixassisted laser desorption/ionization time-of-flight mass spectrometry. Most cleavages predominantly occurred at the C-termini of aspartyl residues. For comparison, weak acid hydrolysis was also performed in boiling water for 20, 40, 60, and 120 min. A 60- min weak acid hydrolysis in boiling water yielded similar results as a 60-min microwave-assisted weak acid hydrolysis at $100^{\circ}C$. These results strongly suggest that microwave irradiation has no notable enhancement effect on acid hydrolysis of proteins and that temperature is the major factor that determines the effectiveness of weak acid hydrolysis.

• 한국수정란이식학회지
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• 제20권3호
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• pp.289-295
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• 2005

본 연구는 돼지에서 스트레스 관련 HSP70 유전자의 다형성과 IVF 수정란의 배발달간의 관련성을 조사하기 위하여 수행되었으며 그 결과는 다음과 같다. HSP70-K1, -K3 및 -K4의 PCR 산물로부터 SSCP 다형성은 각각 다르게 확인되었다. 체외수정란의 분할율 시험에서 HSP70 K1-AA($73.1\%$) 및 K1-AB($62.3\%$) 난할율은 HSP70 K1-BB($49.3\%$)보다 유의적으로 높게 나타났다(p<0.05). 또한 HSP70 K3-AA($72.4\%$) and K3-AB($62.2\%$)형도 HSP70 K3-BB($49.1\%$)형보다 유의적으로 높았다(p<0.05). 돼지품종들과 정자의 HSP70 유전자형에 따른 2-cell 단계까지의 체외수정란 배발달은 유의적 차이를 보였다. Landrace(28.8)와 Duroc(29.8)에서 2세포기까지 발달된 수정란의 수는 Yo.kshire(10.9) 보다 유의적으로 높았다(p<0.05). 또한 HSP70 K4-AB(29.6)형의 2세포기 단계까지의 배발달 수정란 수는 HSP70 K4-AA(10.6)형보다 유의적으로 높았다(p<0.05). 그러나 돼지 품종 및 HSP70 유전자형에 따른 배반포기까지 수정란 배발달 정도는 유의적 차이는 없었다. 이러한 결과들은 돼지 체외수정란의 초기 배발달은 HSP70 유전자형과 품종에 따라서 영향을 받는 것으로 확인되었다.

• 한국수정란이식학회지
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• 제24권3호
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• pp.183-187
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• 2009

In this study, we aimed to determine whether the evaluated markers of cell death could be found at particular developmental stages of normal porcine in vitro fertilization (IVF) embryos. We investigated the characteristics of spontaneous and induced apoptosis during preimplantation development stages of porcine IVF embryos. In experiment 1, to induce apoptosis of porcine IVF embryos, porcine IVF embryos at 22h post insemination were treated at different concentration of actinomycin D (0, 5, 50 and 500 ng/ml in NCSU medium). Treated embryos were incubated at $39^{\circ}C$ in 5% $CO_2$, 5% $O_2$ for 8h, and then washed to NCSU medium and incubated until blastocyst (BL) stage. We examined cleavage rate at 2days and BL development rate at 7days after in vitro culture. A significantly lower rate of cleavage was found in the 500 ng/ml group compared to others (500 ng/ml vs. 0, 5, 50 ng/ml; 27.8 % vs. 50.0%, 41.2%, 35.9%), and BL formation rate in 500 ng/ml was lower than that of others (500 ng/ml vs. 0, 5, 50 ng/ml; 8.0% vs. 12.6%, 11.2%, 12.6%). In experiment 2, to evaluate apoptotic cells, we conducted TUNEL assay based on morphological assessment of nuclei and on detection of specific DNA degradation under fluorescence microscope. This result showed that apoptosis is a normal event during preimplantation development in control group (0 ng/ml actinomycin D). A high number of BL derived control group contained at least one apoptotic cell. Actinomycin D treated BLs responded to the presence of apoptotic inductor by significant decrease in the average number of blastomeres and increase in the incidence of apoptotic cell death. In 500 ng/ml group, the incidence of apoptosis increased at 4-cell stage and later. This result suggested that apoptosis is a process of normal embryonic development and actinomycin D is useful tool for the apoptosis study of porcine preimplantation embryos.