• Journal of Apiculture
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• v.32 no.3
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• pp.181-189
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• 2017

Lysinibacillus fusiformis has been suspected to be a pathogen of Bombus terrestris in Korea since 2008. In this study, we developed the rapid detection method for the L. fusiformis by utilizing the Ultra-rapid PCR. After optimizing of L. fusiformis-specific Ultra-rapid PCR, it can detect the existence of $1.0{\times}10^8$ L. fusiformis-specific DNA molecules in 4 minute and 22 seconds. Even, only 10 molecules could be detected quantitatively using this method. In addition, for the first time, in our knowledge, L. fusiformis was detected using proposed method from bumblebee produced commercially in Korea. Not only in the laboratory but also in the field, L. fusiformis-specific Ultra-rapid PCR would be applied and might be expected as convenient tools at production of bumblebee or inspection for the import and export of bumblebee.

• Clinical and Experimental Reproductive Medicine
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• v.36 no.4
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• pp.265-274
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• 2009

Objective: MicroRNAs (miR) are known to repress target genes at post-transcriptional level and play important roles in development and maturation of cell. However, the expression profiles of miR during ovarian follicle maturation have not been fully elucidated. Here, we designed this study to investigate the expression profiles of miR in oocytes and granulose cells (G-cells) after in vitro culture according to gonadotropins and adding hCG. Methods: Ovaries from 12-day-old mice (C57BL6) were removed and preantral follicles were isolated and cultured in $20\;{\mu}L$-drop of culture media with supplementation of either rFSH, rLH, or rFSH+rLH. After their full maturation, follicles were incubated with rhCG and rEGF. RNA was isolated from oocytes and G-cells, and real-time PCR were performed with primers of miR known to be expressed in the mouse ovary (mmu-miR-16, -miR-27a, -miR-126, -miR-721). Results: FSH+LH group showed the highest ovulation and MII rates among gonadotropin groups. The profiles of miRs in oocytes and G-cells differed according to gonadotropin groups and adding hCG. The profiles of miRs showed divergent changes between oocytes and G-cells. Conclusion: miR expression profiles are altered by gonadotropins and supplementation of hCG during in vitro maturation of murine follicles. Target gene study must be necessary to validate these findings.

• KSBB Journal
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• v.14 no.5
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• pp.593-599
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• 1999

In this study, optimal conditions to infect CD34 positive cells containing hematopoietic stem cells obtained from cord blood and bone marrow were found using two different retroviral vectors expressing human growth hormone (hGH) and $\beta$-galactosidase. CD34 positive cells were successfully infected with recombinant retroviruses only when the CD34 positive cells were co-cultured with packaging cells secreting recombinant retroviruses. To find the highest infection efficiency for the gene transfer, CD34 positive cells from cord blood were co-cultured with packaging cells secreting recombinant retroviruses encoding E. coli lacZ gene. The highest infection efficiency was obtained when CD34 positive cells were cultured for 3 days, and then co-culturing was done for another 2 days. When CD34 positive cells from bone marrow were co-cultured with packaging cells secreting recombinant retroviruses encoding hGH gene, the maximum amount of hGH was also secreted at the same conditions found above, i.e. 3 days of culture and 2 days of co-culture. These results show that there are optimal conditions for the gene transfer into hematopoietic stem cells regardless of sources of target cells or retroviral vectors used to infect.

• Journal of Life Science
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• v.24 no.10
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• pp.1046-1054
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• 2014

Beta-1,3-glucanase is widely used in various biotechnological and industrial processes, with over-production required to enable versatile utilization. We examined the overexpression of ${\beta}$-1,3-glucanase (EXGA) from Aspergillus oryzae using ${\delta}$-sequence-mediated integration. We constructed $pRS{\delta}$-exgA and $pRS{\delta}K$-exgA plasmids for integration of the EXGA gene into various chromosomes of Saccharomyces cerevisiae. These plasmids contain the ADH1 promoter for constitutive expression, a signal sequence (exoinulinase signal sequence [INU1 s.s]) for secretory production, and a ${\delta}$-sequence for integration of ${\beta}$-1,3-glucanase. The $pRS{\delta}$-exgA plasmid was transformed into the S. cerevisiae $BY4742{\Delta}exg1$ strain, and ${\beta}$-1.3-glucanase was stably overexpressed and secreted. Another plasmid, $pRS{\delta}K$-exgA, was introduced into the S. cerevisiae $BY4742{\Delta}exg1$ (YKY082) strain, and overexpression of ${\beta}$-1,3-glucanase was examined by inducible integration under geneticin selection. The activity of ${\beta}$-1,3-glucanase increased in accordance with a rise in the geneticin concentration, with 0.8 mg/ml of geneticin suitable for overexpression of ${\beta}$-1,3-glucanase. Subsequently, $pRS{\delta}K$-exgA was repeatedly transformed for sequential ${\delta}$-integration. The activity of ${\beta}$-1,3-glucanase reached about 0.063 unit/ml/$OD_{600}$, 0.095 unit/ml/$OD_{600}$, 0.131 unit/ml/$OD_{600}$ and 0.165 unit/ml/$OD_{600}$ by the first, second, third, and fourth round of integration, respectively. According to the increase in the activity of ${\beta}$-1,3-glucanase by sequential ${\delta}$-integration, the copy number (integration rate) of the EXGA gene also increased in various chromosomes. These results suggest that recombinant ${\beta}$-1,3-glucanase activity can be sequentially increased by repeated ${\delta}$-sequence integration.

• KSBB Journal
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• v.13 no.2
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• pp.181-186
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• 1998

In order to obtain high-level production of recombinant human thrombopoietin (rhTPO) in insect cell line, HTI-TN-5B1-4 (TN5), conditions for optimal rhTPO expression such as multiplicity of infection (MOI), the cell density at infection, harvesting time and type of culture method as well as growth media were determined. When TN5 cells were cultured as anchorage-dependent state in 60-mm dish, cell density $2\times^6$ cells,MOI of 10 and Garvesting the culture media at 72 hr post-infection wrere the cinditions for highest rh TPO production. High production of rhTPO was also achieved by using EXPRESS FIVE serum free media rather than SF900II serum free media-1. Anchorage-dependent TN5 cells were adapted as a suspension culture when they were grown in the presence of heparin. TN5 cells were successfully cultured at 0.2 L scale in suspension culture without having aggregation. When TN5 cells were cultured as suspension state, cell density of $0.6\times10^6$ cells/mL, MOI of 1 and harvesting the culture media at 72 hr post-infection, gave the highest yield of rhTPO.

• Investigative Magnetic Resonance Imaging
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• v.16 no.1
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• pp.47-54
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• 2012

Purpose : This study was performed to evaluate the characteristics of rat mesenchymal stem cells (RMSCs) transduced with human ferritin gene and investigate $in$ $vitro$ MRI detectability of ferritin-transduced RMSCs. Materials and Methods: The RMSCs expressing both myc-tagged human ferritin heavy chain subunit (myc-FTH) and green fluorescence protein (GFP) were transduced with lentiviurs. Transduced cells were sorted by GFP expression using a fluorescence-activated cell sorter. Myc-FTH and GFP expression in transduced cells were detected by immunofluorescence staining. The cell proliferative ability and viability were assessed by MTT assay. The RMSC surface markers (CD29+/CD45-) were analyzed by flow cytometry. The intracellular iron amount was measured spectrophotometically and the presence of ferritin-iron accumulation was detected by Prussian blue staining. $In$ $vitro$ magnetic resonance imaging (MRI) study of cell phantoms was done on 9.4 T MR scanner to evaluate the feasibility of imaging the ferritin-transduced RMSCs. Results: The myc-FTH and GFP genes were stably transduced into RMSCs. No significant differences were observed in terms of biologic properties in transduced RMSCs compared with non-transduced RMSCs. Ferritin-transduced RMSCs exhibited increased iron accumulation ability and showed significantly lower $T_2$ relaxation time than non-transduced RMSCs. Conclusion: Ferritin gene as MR reporter gene could be used for non-invasive tracking and visualization of therapeutic mesenchymal stem cells by MRI.

• Korean Journal of Microbiology
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• v.49 no.4
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• pp.309-312
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• 2013

Fungal cell walls consist of various glucans and chitin. Fungi produce chitinases for their growth and development. The inky cap, Coprinellus congregatus, produces at least two different chitinases during its life cycle. Chitinase 1 (chi1) is expresses throughout its life cycle while chitinase 2 (chi2) is expressed at the mushroom autolysing phase. The cloned cDNA of chi1 is successfully expressed as a fusion protein with c-myc in Pichia pastoris, and purified by the affinity chromatography. The optimum pH and temperature of Chi1 was pH 8.0 and $35^{\circ}C$, respectively when 4-nitrophenyl N,N',N"-triacetyl-${\beta}$-D-chitotrioside was used as the substrate. The $K_m$ value and $V_{max}$ for the substrate was 0.780 mM and 0.10 OD $min^{-1}unit^{-1}$, respectively. The addition of purified Chi1 resulted in total growth inhibition against several plant pathogenic fungi such as Alternaria alternata, Fusarium graminearum and Trichoderma harzianum at the concentration of 60 ${\mu}g/ml$.

• Proceedings of the Korean Society of Propulsion Engineers Conference
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• 2000.04a
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• pp.4-4
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• 2000

일반적인 액체추진제 로켓엔진의 연소는 분사제트의 미립화, 액적의 증발, 기상 추진제의 혼합, 화학반응 등, 일련의 물리적 과정들로 이루어지고, 여기서 특성속도 효율은 크게 분사특성 및 연소의 두 단계에서 결정되게 된다. 액체추진제 로켓엔진에 사용되는 여러 분무형태 중, 동축형 분사기에서는 액상과 기상 제트의 운동량 차에 의해 미립화가 이루어지며, 분무 액적들의 전개와 더불어 분사기 출구를 포함한 전 영역에서 연소가 발생되므로 매우 복잡한 물리적 특성들을 포함하게 된다. 본 연구에서는 기상 연료-액상 산화제의 동축형 분무연소를 JANNAF의 방법을 사용하여 수식화 하였으며, 이를 바탕으로 분무특성과 연소성능 예측을 위한 프로그램을 작성, 분사조건에 의한 분무특성과 그에 따른 연소성능을 계산하였다. 연속, 운동량, 에너지 및 혼합비 방정식의 지배방정식들을 바탕으로 기상 유동을 수식화 하였으며, 별도로 액적의 소산 및 연소과정을 모사하기 위한 별도의 수식들이 추가되었고, 이 식들을 결합하여 액적의 크기, 분포를 포함하는 액체 제트의 미립화 정도를 공간적으로 계산하였다. 미립화 모델의 검증을 위하여 계산 결과를 Reitz의 실험과 Giridharan의 모델 등과 비교하였으며 잘 일치하는 경향을 나타내었다. 또한 동축형 분사기에서의 분무 특성을 예측하기 위해 액체 산소, 기체 수소를 추진제 조합으로 하는 동축형 분무 연소장에서의 제트 길이, 액적의 크기, 액체 제트의 속도를 계산하였다. 계산 결과 액체 제트의 접촉길이는 분사공의 지름이 증가할수록 웨버수가 증가되므로 짧아지는 것으로 관찰되었으며 액적의 크기도 분사공의 지름이 증가할수록 작아지는 경향을 나타내었다. 액체 제트의 속도는 처음에는 일정하게 유지되다가 운동량을 보존하기 위해 가스로부터 운동량을 받아 점차 가속되어지는 것으로 나타났다.본 규격은 키, 총장, 어깨길이, 등길이, 머리길이, 머리둘레, 진동둘레, 목둘레, 가슴둘레, 허리둘레, 배둘레, 엉덩이둘레, 앞품, 뒤품, drop치를 포함하고 있고, 각 규격에서 호칭간 치수 간격도 함께 제시하고 있다. 본 연구 결과에서 보듯, 현행 8규격의 무진복의 각 호칭간 적정 허용범위를 고려해 합리적인 치수체계를 정립한다면 치수에 대한 적합도가 상당히 증가할 뿐 아니라 생산비용도 상당히 감축할 것으로 생각된다.나타났다. 4) 호감적 서비스능력 차원에서 세 독립변수간에 유의한 3원 상호작용이 존재하는 것으로 나타나( $F_{2,228}$=15.62, P<.001) 20대에 적합한 의복 착용시( $F_{2,228}$=3.98, P<.05)와 60대에 적합한 의복 착용시( $F_{2,228}$=16.55, P<.001) 점포유형과 격식차림간에는 유의한 상호작용이 존재하는 것으로 나타났다. 5) 호감을 구성하는 세 요인들이 구매의도에 미치는 영향을 조사한 결과 호감적 인상차원은 29%(P<.001), 호감적 서비스능력차원은 6%(P<.001)의 구매의도를 설명해 주는 것으로 나타났다. 본 연구결과 노년 소비자에게 호감을 주는 판매원의 외모는 구매의도에 영향을 주어 실버의류산업의 이익증대와 밀접한 연관을 갖는 서비스품질의 중요한 요인으로 밝혀졌다.중요한 요인으로 밝혀졌다.로운 단백질 EPSPS가 다른 여러 식물에 이미 존재하고 있는 단백질로서 우리가 이미 이러한 식품을 섭취할 때 이 단백질도 같이 섭취해오고 있었다는 점, 둘째. 이 단백질이 소화액 분해 실험에서 짧은 시간내에 분해가 되었다는 점, 셋째. 재조합 된 콩과 자연 콩이 성분 분석에서 차이를 나타내지 않았다는 점, 네 번째. 쥐를 통한 다양섭취 실험에서 아무런 이상 반응이 없었

• Journal of Life Science
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• v.8 no.3
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• pp.263-271
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• 1998

One of the better-characterized transcription factor of E. coli is the cAMP receptor protein(CRP) and the CRP binds cAMP and DNA. The cAMP-CRP complex is involved in regulation of many genes at bacteria. The cAMP-CRP regulatory element represents, in some respects, a global regulatory network. The aim of this work was to study the structure and the mechanisms controlling the expression of CRP in Serratia marcescens. We have been get 5 different clones from Serratia which stimulated the cells to use maltose as a sole carbon source in E. coli TP2139. The crp gene clone, pCKB12, was confirmed by Southern hybridization with E. coli crp gene. The location of the crp gene was determined by construction subclones carrying various portions of pCKB12. To investigate the potential role of CRP in E. coli, lacZ fused plasmids were constructed and investigated the ${\beta}$-galactosidase activity of the fused plasmid. The Serratiamarcescens cAMP receptor protein can substitute the E. coli CRP in transcriptional activation at the lacZ gene. These results suggest that Serratia marcescens cAMP receptor protein complex functions to regulate several promoters in E. coli.

• Korean Journal of Animal Reproduction
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• v.20 no.4
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• pp.371-378
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• 1997

Human lactoferrin (hLF) was expressed in the mammary gland of transgenic mice. Expresion of hLF was achieved by palcing its cDNA under the control of bovine $\beta$-casein gene. To improve the hLF expression level, two artificial introns were introduced into the expression vector. One intron is a hybrid-splice consisting of bovine $\beta$ casein intron 1 and rabbit $\beta$-casem intron II. The other intron is a DNA fragment spanning intron 8 of bovine $\beta$ casein gene. Trans sgenic mice were developed which expressed hLF in their milk. Twenty lines of transgenic mice were produced. hLF was present in the milk at concentrations of 1 ~ 200 ${\mu}\textrm{g}$ / ml. hLF RNA was only detected in the mammary gland of transgenic mice. The expressed RNA was cor r rectly spliced at the exon /intron junctions. To generate transgenic cows secreting active hLF in their milk, we transferred the DNA-injected bovine embryos to recipient heifers by surgical a and non-surgical methods out of 68 embryos transferred to 51 recipients by surgical or non-surgical method, 7 calves were normally born. Effect of embryo quality of DNA-injected blastocysts on pregnancy rate after transfer was investig a ated. Higher pregnancy rate of (38.9%) DNA-injected embryos was shown in excellent embryos. Pregnancy rates in the groups of good a and fair embryos were 15.4 and 14.3%, respectively. Effect of culture period of DNA-injected b bovine embryos on pregnancy rate after transfer was investigated. When Day-6 blastocysts of cuI ture were transferred, there was no pregnancy. Pregnancy rates of Day-7 and -8 blastocysts were 28.6 and 33.3%, respectively. There was no difference on pregnancy rate between Day-7 a and -8 bovine blastocysts after DNA injection. Thus, we established the techniques for transfer a and culture of DNA-injected bovine embryos. In a addition, factors affecting the pregnancy rate of DNA-injected embryos after transfer were investigated .