• Journal of Sericultural and Entomological Science
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• v.52 no.1
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• pp.25-32
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• 2014

We produced the transgenic silkworm that expressed the enhanced blue fluorescent protein (EBFP) in the cocoon of silkworms. The EBFP fusion protein, each with N- and C-terminal sequences of the fibroin H-chain, was designed to be secreted into the lumen of the posterior silk glands. The expression of the EBFP/H-chain fusion gene was regulated by the fibroin H-chain promoter. The use of the $3{\times}P3$-driven DsRed2 cDNA as a marker allowed us to rapidly distinguish transgenic silkworm. A mixture of the donor and helper vector was micro-injected into 300 eggs of silkworms, Baegokjam. We obtained 5 broods. The cocoon displayed blue fluorescence, proving that the fusion protein was present in the cocoon. Also, the presence of fusion proteins in cocoons was demonstrated by SDS-PAGE and western blot analysis. Accordingly, we suggest that the EBFP fluorescence silk will enable the production of the silk-based biomaterials.

• Journal of Life Science
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• v.19 no.6
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• pp.809-817
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• 2009

The phenotypes associated with a gene function are often the best clue to its role in the plant. Transgenic plants ectopically expressing or suppressing a gene can provide useful information related to the gene function. In this study, we constructed three vectors - pFGL571, pFGL846 and pFGL847 - for the Agrobacterium-mediated ectopic expression of plant genes using pPZP211 and modified CaMV 35S, UBQ3 or UBQ10 promoters. The three vectors have several merits such as small size, high copy in bacteria, enough restriction enzyme sites in multi cloning sites and nucleotide sequence information. Analysis of transgenic plants containing GUS or sGFP reporter genes under the control of modified CaMV 35S, UBQ3 or UBQI0 promoter revealed that all of the three promoters showed high activities during most developmental stages after germination and in floral organs. Furthermore, we generated a RNAi module vector, pFGL727, to suppress plant gene expressions and confirmed that pFGL727 is useful for the suppression of a gene expression using rice transgenic plants. Taken together, our new vectors would be very useful for the ectopic expression or the suppression of plant genes.

• Proceedings of the Korean Society of Developmental Biology Conference
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• 2003.10a
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• pp.66-66
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• 2003

Protein tyrosine kinases는 표적단백질의 tyrosine 잔기를 인산화하는 효소로서 다양한 종류의 성장인자, peptide 호르몬, cytokine 수용체 하위의 세포 내 신호전달에 관여한다. Non-receptor tyrosine kinase의 일종인 c-Src는 세포막에서 발생한 ligand-receptor 상호작용 하위의 신호전달에서 중요한 역할을 하며 C-terminal Src kinase (Csk)는 Src kinase의 C-terminal tyrosine 잔기를 인산화시켜 Src kinase의 활성을 저해한다. 이러한 Src-Csk loop를 통한 세포 내 신호전달과정은 세포의 증식과 분화, 사멸 조절에 중요한 기능을 갖지만 정소의 발생과 분화 과정에서 Src-Csk loop의 발현 및 정자형성 과정에서의 기능은 밝혀지지 않았다. 본 연구에서는 생쥐 정소에서 출생 후 성적 성숙과정에서 Csk의 발현과 Src kinase 활성의 변동을 조사하였다. Csk mRNA 발현은 생 후 2주령 이하의 미성숙 정소에서 다량으로 발현되었고 사춘기 정소 이후에는 오히려 감소하였다. Csk 단백질의 발현 양상은 mRNA 발현양상과 일치하였다. c-Src kinase 활성은 생 후 2주에 급격히 증가하고 이 후 4주령에서 감소하다가 성체 (8주령)에서 다시 증가하여 가장 높았다. 성체 조직의 Csk 단백질 현존량이 미성숙 개체보다 적은 반면 Src kinase 활성은 가장 높아 Csk 발현의 감소는 Src kinase 활성을 증가하는 것으로 사료된다. 면역조직화학방법으로 정소 조직 내 Csk의 발현양상을 조사한 결과 Leydig cell, Sertoli cell, germ cell 등 도처에서 발현되었으며 Sertoli cell 에서의 발현은 세정관 상피의 구성에 따른 차이가 확인되었다. 성체의 세정관 내에서는 감수분열 이후의 정세포(spermatid)를 감싸고 있는 Sertoli cell의 강소측에서 강한 Csk 활성이 검출되어 생식세포의 분화과정 동안 세정관 상피의 조직재구성에 관여하는 것으로 사료된다. Leydig cell에서의 발현은 생후 1주령까지는 미미하였으나 이후 2주령 이후에는 다량으로 발현함이 확인되어 adult type Leydig cell에서 진행되는 steroidogenesis와의 관련성을 추측할 수 있다. 미성숙 정소로부터 분리한 Sertoli cell-enriched culture에 200 nM testosterone을 처리하였을 때 Csk mRNA의 발현의 증가를 확인할 수 있었으므로 androgen에 의한 Sertoli cell의 분화과정에 Csk가 관여하고 있음을 알 수 있다. 결론적으로 성적 성숙에 따른 생쥐 정소 내 Src-Csk loop의 발현과 Src kinase 활성의 변동은 정소 내 간충조직, 세정관 상피의 증식 및 기능적 분화 과정을 매개하는 생리적 활성분자 수용체 하위의 신호전달 과정에 Src-Csk loop에 의한 조절가능성을 확인할 수 있었다.

• Development and Reproduction
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• v.5 no.2
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• pp.131-136
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• 2001

The developmental profiles of rpr, grim, dcp-1, diapl, diap2 transcripts, which were involved in programmed cell death, were analyzed using competitive RT-PCR in whole animals during Drosophila development. The fluctuation patterns of transcript levels of the apoptotic initiators(rpr and grim) were similar to those of the ecdysone titer in Drosophila life cycle. The transcript of dcp-1, which is considered as effector caspase, was expressed strong1y at early embryo and female adult stages. However, the transcript levels of anti-apoptotic factors diap1 and diap2, showed the reverse pattern comparing with those of apoptotic factors(rpr and grim). Also, the transcript levels of rpr, diap2 and dcp-1 were quantified in the salivary glands and wing discs dissected from the wandering late third instar larva. The transcript levels of rpr and diap2 were changed reversely each other in both tissues from wandering stage to puparium formation. These results suggest that the expressions of cell death related genes are regulated by the ecdysone signals during normal development.

• Development and Reproduction
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• v.11 no.1
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• pp.21-29
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• 2007

Previously, we found progesterone receptor membrane component 2 (pgrmc2) was highly expressed in germinal vesicle (GV) stage oocytes. The present study was conducted to characterize the expression of pgrmc2, as well as pgrmc1, in the mouse gonads and embryos according to their developmental stages. We found that these membrane components were expressed in ovaries, testes, and embryos at various developmental stages in addition to oocytes. Progesterone-3-O-carboxymethyl oxime-BSA-fluorescein isothiocyanate (P4-BSA-FITC) was applied to visualize the presence of the progesterone receptor on mouse oocyte membrane, and we confirmed that immobilized progesterone is localized at surface of the oocyte. This is, at our knowledge, the first report regarding the expression of membrane component of progesterone receptor in the mouse oocytes, embryos, and gonads. The function and signal transduction pathway of progesterone receptor membrane components in oocytes requires further studies.

• Proceedings of the Korean Society of Applied Pharmacology
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• 1993.04a
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• pp.138-138
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• 1993

인슈린 유사체를 유전공학적 방법으로 생산하여 새로운 당뇨병 치료제로써의 가능성을 타진한다. 인슈린 B 사슬의 C 말단 아미노산 codon을 threonine 대신을 methionine을 지령하도록 하고 여기에 인슈린 A사슬을 지령하는 염기를 바로 부착시켜, 이를 대장균에서 발현시키므로써 외사슬 인슈린 선구체를 제조하고, 이를 분리 정제한 다음 취화브롬 으로 절단하므로서 ($B^{30}$-homoserine) 인슈린을 제조한다. 1. 외사슬 인슈린 전구체 유전자를 합성한 후 대장균의 발현 운반체내 e-galactosidase 유전자와 융합시켜 도입하므로서 재조합된 융합단백질을 생산하였다. 2. 재조합 대장균을 발효한 후 urea 융합단백질을 분리하고 DEAE-Sephacel과 Sephadex G-200을 이용하여 순수 정제하였다.

• The Microorganisms and Industry
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• v.17 no.2
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• pp.2-10
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• 1991

식물체에서 일어나는 유전자 발현의 광조절 현상은 매우 복잡하다. 이는 각 유전자들이 조직에 따라 질적, 양적, 시간적 측면에서 빛에 대한 반응에 커다란 차이점을 보이고 있기 때문이다. 따라서 식물체를 대상으로 광조절 기작을 연구하는 데는 현상의 복합성과 실험과정의 기술적, 시간적, 경제적 제약이 많기 때문에 이에 관한 연구가 아직 초보적 수준에 머물고 있다. 광합성 세균 중에서 purple bacteria나 green bacteria와는 달리 식물성 광합성을 하며, 식물체보다 세포구조가 훨씬 간단한 cyanobacteria는 유전자 발현의 광조절 기작을 연구하는데 간단, 명료한 'model'로서 기대되는 바 크다. 따라서 이 글에서는 cyanobacteria의 광계와 광합성 색소인 chlorophyll의 생합성 과정을 중심으로 광조절 현상에 대한 최근 연구 동향과 전망을 살펴보고자 한다.

• Proceedings of the Zoological Society Korea Conference
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• 1994.10a
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• pp.263.2-263
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• 1994

• Development and Reproduction
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• v.6 no.2
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• pp.123-129
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• 2002

Leptin, a product of the obese gene, is associated not only with obesity but also with female reproductive function, but it has not yet been ascertained whether leptin acts directly on the ovary or indirectly via the hypothalamus-pituitary pathway. Therefore, the object of this study was to investigate the expession of leptin and its receptor in the rat ovary by immunohistochemistry and RT-PCR during the estrous cycle. Immunohistochemistry results showed that leptin was stained in the theca cells and in part of granulosa cells in atretic follicles, whereas leptin receptor was localized in the interstitial cells and ova in preantral follicies. In particular, leptin and its receptor in atretic follicles displayed more intensive staining compared to those in normal follicles. During the estrous cycle, the mRNA expression of leptin and its receptor in the ovary was detected by RT-PCR and estradiol, progesterone, and leptin levels in the serum was measured by ELISA. The leptin level in the serum on metestrous phase was significantly higher than that on estrous phase. Similar to leptin level, progesterone level increased on metestrous phase. Leptin mRNA was not detected throughout the estrous cycle, whereas leptin receptor mRNA was expressed on all phases of estrous cycle excepting the diestrous phase. These results suggest that leptin might be directly involved in the regulation of ovarian function in rat.

• Clinical and Experimental Pediatrics
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• v.46 no.4
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• pp.376-381
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• 2003

Purpose : This study aimed to demonstrate the possible pathogenesis of granulopoiesis in patients of Kawasaki disease(KD) using quantitative analysis of G-CSF, GM-CSF and their CSFr. Methods : The plasma levels of G-CSF, GM-CSF, G-CSFr and GM-CSFr were studied in 14 patients in the acute phase of KD; 13 children with normal peripheral white blood cell counts were used as the normal control group. The plasma concentration of G-CSF, GM-CSF were analyzed by ELISA. The G-CSFr and GM-CSFr on the peripheral granulocytes were analyzed by a quantitative flow cytometric assay and QuantiBRITE, and the quantitative changes of receptors which did not combine with G-CSF and GM-CSF were measured. Results : The total number of leukocytes in KD was similar to normal control group, but the leukocytes increased according to the number of neutrophils. The plasma concentration of G-CSF were decreased similar to normal control group(P=0.133), but that of GM-CSF decreased more than the normal control group(P=0.227). The quantity of G-CSFr, GM-CSFr were revealed to be no less than the normal control(P=0.721, P=0.912). After incubation with excessive G-CSF, the expressed G-CSFr on the neutrophils were decreased in both groups(P=0.554). The quantities of expressions of GM-CSFr on the neutrophil after incubation with the excessive GM-CSF were always increased in both groups(P=0.255). The amount of GM-CSFr of neutrophils are in proportion to total white blood cells (r=0.788, P=0.035), but it wasn't in the case of KD(P=0.644). Conclusion : The leukocytosis in KD that mediated by increasing neutrophil was not correlated with the plasma concentrations of G-CSF and GM-CSF, and the amount of expression of G-CSFr and GM-CSFr on granulocyte. It is possible that the reduction of concentration of GM-CSF results by increasing the active GM-CSFr.