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| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.author | Wei-Yuan Tsai | en |
| dc.contributor.author | 蔡維原 | zh_TW |
| dc.date.accessioned | 2021-07-01T08:13:11Z | - |
| dc.date.available | 2021-07-01T08:13:11Z | - |
| dc.date.issued | 2003 | |
| dc.identifier.citation | Eduardo Marb?n. Cardiac channelopathies. Nature 415, 213-218 (2002). Efrat S., Fusco-DeMane D., Lemberg H., al Emran O and Wang X. Conditional transformation of a pancreatic beta-cell line derived from transgenic mice expressing a tetracycline-regulated oncogene. Proceedings of the National Academy of Sciences of the United States ofAmerica 92, 3576-35 80 (1995). Fahmy R.G. and Khachigian L.M. Antisense Egr-1 RNA driven by the CMV promoter is an inhibitor of vascular smooth muscle cell proliferation and regrowth after injury. Journal of Cellular Biochemistry84, 575-82 (2002). Furth P., St Onge L., Boger G, Gruss P., Gossen M., Kistner A., Bujard H., and Hennighausen L. Temporal control of gene expression in transgenic mice by tetracycline-responsive promoter. Proceedings of the National Academy of Sciences of the United States of America 91,9302-9306 (1994). Grill M.A., Bales M.A., Fought A.N., Rosburg K.C., Munger S.J., and Antin P.B. Tetracycline-inducible system for skeletal muscle-specificgene expression in transgenic mice. Transgenic Research 12, 33-43(2003). Gossen M. and Bujard H., Tight control of gene expression in mammalian cells by tetracycline-responsive promoters._Proceedings of the National Academy of Sciences of the United States of America 89, 5547-5551(1992). Huang C. J., Tu C. T., Hsiao C. D., Hsieh F. J., and Tsai H. J., Germ-line transmission of a myocardium-specific GFP transgenic reveals critical regulatory elements in the cardiac myosin light chain 2 promoter of zebrafish. Developmental Dynamics 228 (2003). Sehnert. A. J., Huq A., Weinstein B.M., Walker C., Fishman M., and Stainier D.Y.R. Cardiac troponin T is essential in sarcomere assembly and cardiac contractility. Nature Genetic 31, 106-110 (2002). Sehnert. A.J., and Stainier D.Y.R. A window to the heart: can zebrafish mutants help us understand heart disease in human Trends in Genetics18, 49 1-494 (2002). Towbin J.A., and Bowles N.E. The failing heart. Nature 415, 227-233 (2002). Wang X., Li M.X., and Sykes B.D., Structure of the regulatory N-domainof human cardiac Troponin C in complex with human cardiac Troponin I147-163 and bepridil. Journal of Biological Chemistry 277, 31124-31133(2002). Wargelius A., Ellingsen S., and Fjose J. Double-stranded RNA induces specific developmental defects in zebrafish embryos. Biochemical and Biophysical Research Communications 263, 156-161 (1999). Zhixing Z., Cao Y., Li M., and Meng A. Double-stranded RNA injection Preduces nonspecific defects in zebrafish. Developmental Biology 229,215-223 (2001). | |
| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/75433 | - |
| dc.description.abstract | Tet System 是由 Gossen 跟Bujard 在 Escherichia coli發現的 transcriptional repression system 所發展而來，藉由 tetracycline 跟 tTA ( transcriptional repressor ）結合後，可抑制欲研究基因的表現。而後 Gossen 等人又將 tTA 結合VP16 ( transcriptional activator）而成 rtTA ，使得在 tetracycline 存在的環境下，rtTA 會跟 tetracycline 結合並開敔特定基因的表現 ( tet-on system ）。在我們的實驗中，利用 tet-on system 配合斑馬魚心臟專一表現基因（ cardiac myosin light chain ）?動子，在斑馬魚心臟專一表現 cardiac Troponin C（cTnC）。cTnC 是一個鈣離子結合蛋白，在cTnC 和鈣離子結合後，actin可以跟 myosin 結合使得肌纖維產生收縮。我們架構了一個心臟專一表現的雙向?動子，用以同時驅動EGFP報導基因以及 cTnC-antisense 在心臟專一表現，利用 antisense 抑制內生性基因表現，來研究 cTnC 的基因功能。 在基因轉殖魚胚胎發育到 12 小時，添加入 tetracycline 衍生物 doxycycline ，可以在 12 小時後看到綠色螢光在心臟專一表現。在 48 小時大的魚，可以觀察到 cTnC-antisense 的心跳速率顯著的較控制組來的快（ cTnC – antisense l66.8 每分鐘；控制組 137.6 每分鐘） ; 但是在 6 天大的魚中， cTnC-antisense 的心跳數卻是明顯的慢於控制組（cTnC-antisense 190.9 每分鐘；控制組219.5 每分鐘）。當進行 cTnC morpholino 注射實驗時，在 1.15ng 劑量注射過的魚中，會出現心室不會跳動的性狀；而在 0.575ng 劑量注射的魚中，並未出現心室停止跳動的性狀，跟控制組比較起來，心跳數有些微變快的性狀。藉由 cTnC-antisense 所產生心臟異常性狀跟藉由較低劑量 cTnC morpho1ino 所產生的變異相似。藉由這樣的證據，可以支援藉由 tetracycline regulatory system 所驅動的可調控的基因表現系統在斑馬魚中可以使用，同時也提供另一種研究特定基因的實驗方式。 | zh_TW |
| dc.description.abstract | The tetracycline-controlled transcription system provides a convenient method to study stage-depended gene expression. Here, we report an inducible tet-on system to study the zebrafish cardiac troponin C (cTnC), a calcium binding protein. We constructed a bi-directional vector, in which GFP and antisense of cTnC were driven by a cardiac specific promoter, cmlc2. When 12-hpf embryos were treated with 1 μg/ml doxycycline, we found that GFP was specifically expressed in the hearts of embryos after induction for 12 hr. We also found that the hearts were slightly bigger than that of wild-type embryos and the heart-beating rate was little faster than wild-type (166.8 vs 137.6 per minutes). When cTnC-morpholino (cTnC-MO) was injected into one-celled fertilized eggs, the 1.15ng-injected embryos showed a silent ventricle syndrome. No silent ventricle occurred in the 0.575ng-injected embryos and instead the heart —beating rates were slightly faster than those of wild-type. The heart defects were caused by cTnC knock-down due to cTnC-antisense induction was similar to those caused by the lower dosage treatment of cTnC-MO. These evidences strongly suggest that this inducible system may work on model fish so that it provides an approach to study gene function at any desired developmental stages. | en |
| dc.description.provenance | Made available in DSpace on 2021-07-01T08:13:11Z (GMT). No. of bitstreams: 0 Previous issue date: 2003 | en |
| dc.description.tableofcontents | 中文摘要---------------------------------------------------------1 英文摘要--------------------------------------------------------3 壹．前言---------------------------------------------------------4 貳．材料與方法---------------------------------------------------------7 參．結果---------------------------------------------------------l2 肆．討論---------------------------------------------------------l5 伍．參考資料 ---------------------------------------------------------l8 圖表---------------------------------------------------------21 附錄 ---------------------------------------------------------27 | |
| dc.language.iso | zh-TW | |
| dc.title | 利用tetracycline regulatory system 表現 cardiac Troponin C | zh_TW |
| dc.title | Using tetracyc1ine-controlled transcriptiona1 system to express zebrafish cardiac troponin C | en |
| dc.date.schoolyear | 91-2 | |
| dc.description.degree | 碩士 | |
| dc.relation.page | 52 | |
| dc.rights.note | 未授權 | |
| dc.contributor.author-dept | 生命科學院 | zh_TW |
| dc.contributor.author-dept | 漁業科學研究所 | zh_TW |
| 顯示於系所單位： | 漁業科學研究所 | |
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