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2006 諾貝爾生醫獎 RNA Interference

2006 諾貝爾生醫獎 RNA Interference. 組別 : 第一組 組長 :B9602026 林旻暭 組員 :B9602006 江柏興 B9602009 余志恆 B9602034 胡家祥 B9602042 張晢鏞. 歷史年表. 1990- 發現紫色牽牛花的 co-supression 現象。 1994- co-supression 被認為是在後轉錄時期發生，因此 以 Post-transcriptional Gene Silencing 稱之。

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2006 諾貝爾生醫獎 RNA Interference

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  1. 2006諾貝爾生醫獎RNA Interference 組別:第一組 組長:B9602026林旻暭 組員:B9602006江柏興 B9602009余志恆 B9602034胡家祥 B9602042張晢鏞

  2. 歷史年表 • 1990-發現紫色牽牛花的co-supression現象。 • 1994- co-supression被認為是在後轉錄時期發生，因此 以Post-transcriptional Gene Silencing稱之。 • 1995- Gao 和 Kemphues將sense及antisense RNA注入 Caenorhabditis elegans(秀麗隱桿線蟲)體內。 • 1998- Andrew Z. Fire和Craig C. Mello將dsRNA注入 C. elegans發現了有效的gene silencing 。 • 2000-RNAi的路徑被提出。 • 2001-發現siRNA和dicer 。 • 2002-RNAi開始應用在HIV的治療上。

  3. 1990紫色牽牛花實驗

  4. 1998線蟲實驗 Caenorhabditis elegans(秀麗隱桿線蟲)

  5. 1.將unc-22 gene注入C. elegans

  6. 2.利用GFP篩選

  7. 3.將mex-3 RNA注入C. elegans胚胎

  8. 反義寡核酸分子 正常轉譯-基因表現 核酸酵素 miRNA RNAi 作用機制 siRNA

  9. RNAi 作用機制 – Dicer RNase III domain amino-terminal helicase domain connector helix PAZ domain

  10. RISC

  11. RNAi的信號擴增系統 • 被RISC切斷的mRNA片段可以被RdRp(RNA-Dependent RNA polymerase)附著，並進行複製，形成一dsRNA。Dicer酶再對dsRNA進行切割，造出新的雙鏈siRNA。如此不斷循環，便可達到信號擴增的效果。

  12. Vector-Based siRNA Technology • The source of siRNA • synthetic RNA duplexes( two unmodified 21-oligonucleotide) • manually designed • siRNA Design Guidelines • Find 21 nt sequences in the target mRNA that begin with an AA dinucleotide • Select 2-4 target sequences • Design appropriate controls

  13. DNA → siRNA sense siRNA sequence antisense siRNA sequence short spacer 5-6 T's EcoR I and Apa I restriction sites Procedure for siRNA Hairpin Design

  14. U6 ：RNA pol Ⅲpromoter RNA Polymerase Ⅲ expresses relatively large amounts of small RNAs in mammalian cells terminates encountering a string of 4-5 uridines pSilencer™siRNA Exression Vector

  15. Application Hepatitis B virus (HBV) cure Traditional：anti-viral drugs Genetically modified adenovirus Food：peanuts p34, causing65%of soybean allergies) muscle-derived stem cells (MDSCs) to become bone-forming cells MyoD1(myogenesis ) & Smad6 (osteogenetic Treating musculoskeletal diseases and injuries Cure for AIDS

  16. Strategy viral mRNAs rev gag gene( Gag precursor protein→p24: core) Cellular mRNAs receptor: CD4 coreceptor: CC chemokine receptor 5 high error rate of HIV siRNA : a change in even 1 bp drastically lowers the potency of siRNA To cell or virus??

  17. 資料來源 • 維基百科-RNA干擾http://zh.wikipedia.org/w/index.php?title=RNAi&variant=zh-tw • 維基百科-安德魯法厄http://zh.wikipedia.org/w/index.php?title=%E5%AE%89%E5%BE%B7%E9%B2%81%C2%B7%E6%B3%95%E5%8E%84&variant=zh-tw • 維基百科-克雷格·梅洛http://zh.wikipedia.org/w/index.php?title=%E5%85%8B%E9%9B%B7%E6%A0%BC%C2%B7%E6%A2%85%E6%B4%9B&variant=zh-tw • 維基百科-轉座子Transposonhttp://zh.wikipedia.org/w/index.php?title=%E8%BD%AC%E5%BA%A7%E5%AD%90&variant=zh-tw • RNAi相關實驗http://www.ambion.com/techlib/resources/RNAi/overview/index.htmlhttp://faculty.virginia.edu/mammgenetics/805-2ndX03.htmlhttp://www.mgm.ufl.edu/faculty/rrenne.htmhttp://bi.snu.ac.kr/ProMiR/microRNA.htmhttp://www.answers.com/topic/sirna-structure2-jpg

  18. RNAi動畫http://www.nature.com/focus/rnai/animations/index.htmlRNAi動畫http://www.nature.com/focus/rnai/animations/index.html • 2006諾貝爾生醫獎(官方網站)http://nobelprize.org/nobel_prizes/medicine/laureates/2006/index.html(Advanced information)http://nobelprize.org/nobel_prizes/medicine/laureates/2006/adv.html • RNAi應用 • RNA Interference — A New Weapon against HIV and Beyondhttp://content.nejm.org/cgi/content/full/347/17/1364 • siRNA Design Guidelineshttp://www.ambion.com/techlib/tb/tb_506.html • siRNA Technologyhttp://www.genscript.com/siRNA_technology.html • RNAi: Silencing never sounded betterhttp://www.nature.com/nmeth/journal/v1/n1/full/nmeth1004-79.html • Fixing Food http://www.sciam.com/article.cfm?id=fixing-food • GM viruses may help cure Hepatitis B infections http://www.scidev.net/News/index.cfm?fuseaction=readNews&itemid=3164&language=1 • Study Finds Possible Targets for H.I.V. Drugs http://www.nytimes.com/2008/01/11/science/11hiv.html?ex=1357794000&en=66e26c43cabac92c&ei=5124&partner=permalink&exprod=permalink • More siRNA Vectors for RNA Interferencehttp://www.ambion.com/techlib/tn/95/952.html

  19. (文書資料) • 當代醫學 第三十三卷 第十二期 72~76 • 張璐璐、楊樹東、張季平，2006，何為「核糖核酸干擾（RNAi）」及其在醫學上之新用途，當代醫學，398：1018~1019 • 生物化學與生物物理的進展 2003; 30 (5): 702~705 • microRNA Brooker, Widmaier, Graham, Stiling, BIOLOGY, p273~274 • International Journal of Information Technology Vol. 11, No. 8, 2005 • Ambros, V. (2004) The functions of animal microRNAs. Nature 431, 350~355 • (資料閱讀) • Cold Spring Harbor Laboratory http://www.cshl.edu/ • RISC http://www.uky.edu/~gtang2/tang-area_files/chapter2.pdf • Hopes http://www.stanford.edu/group/hopes/treatmts/pbuildup/h2.html • Invitrogen http://www.invitrogen.com/content.cfm?pageid=10088 • (圖片來源) • hkmargie.blogspot.com/2007_02_01_archive.html • hkmargie.blogspot.com/2007_02_01_archive.html

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