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| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 林峰輝 | |
| dc.contributor.author | Yi-Cheng Tseng | en |
| dc.contributor.author | 曾意誠 | zh_TW |
| dc.date.accessioned | 2021-06-13T03:16:53Z | - |
| dc.date.available | 2006-07-31 | |
| dc.date.copyright | 2006-07-31 | |
| dc.date.issued | 2006 | |
| dc.date.submitted | 2006-07-28 | |
| dc.identifier.citation | 1. W. A. Kues, H. Niemann, The contribution of farm animals to human health, TRENDS in Biotechnology 22(6), 286-294, 2004.
2. 沈朋志，動物基因轉殖，科學發展，372期，2003年12月，24-29。 3. R. F. Weaver, Molecular Biology, Third edition , 787-792, 2005. 4. 楊卓真，應用精子載體法產製轉基因雞之可行性，2002年六月，碩士論文，台灣大學，台北市。 5. S. G. Lillico, Transgenic chickens as bioreactors for protein-based drugs. Drugs Discovery Today, 10(3) , 191-196, 2005. 6. R. Ivarie, Avian transgenesis: progress towards the promise, TRENDS in Biotechnology 21(1), 14-19, 2003. 7. Petitte, J.N. et al. Production of somatic and germline chimeras in the chicken by transfer of early blastodermal cells. Development 108, 185-189, 1990. 8. Fraser, R.A. et al. Efficient incorporation of transfected blastodermal cells into chimeric chicken embryos. Int. J. Dev. Biol. 37, 381-385, 1993. 9. Carsience, R.S. et al. Germline chimeric chickens from dispersed donor blastodermal cells and compromised recipient embryos. Development 117, 1993, 669-675 10. 馬春祥、吳和光、鄭登貴。家禽之生殖。1997。初版。國立編譯館。台北市。 11. M.N.V. Ravi kumar, U.Bakowsky, C.M Lehr, Preparation and characterization of cationic PLGA nanospheres as DNA carriers, Biomaterials 25, 1771-1777, 2004. 12. M.I. Baraton, Synthesis, functionalization and surface treatment of nanoparticles, American scientific publishers, first editon, USA, 233-256, 2003. 13. K.S. Soppimath, T.M. Aminabhavi, A.R. Kulkarni, W.E. Rndzinski, Biodegradable polymeric nanoparticles as drug delivery devices, Journal of Control Release 70, 1-20, 2001. 14. G. Borchard, Chitosans for gene delivery, Advanced drug delivery review 52, 145-150, 2001. 15. T. Dastan, K. Turan, In vitro characterization and delivery of Chitosan-DNA microparticles into mammalian cells, J Pharm Pharmacent Sol. 7(2), 205-214, 2004. 16. K.Y.Lee, S.Y.Jeong, Preparation of Chitosan self-aggregates as a gene delivery system , Journal of Controlled release 51, 213-220, 1998. 17. 李家成。幾丁聚醣/DNA奈米微粒於基因轉殖雞之應用。2005年。碩士論文。台灣大學，台北市。 18. 陳貽華。 磷鈣奈米微粒作為非病毒基因遞送載體研究。 2004年。 碩士論文。 台灣大學，台北市。 19. Tejraj M. Aminabhavi, Sunil A. Agnihotri, Recent advances on chitosan-based micro- and nanoparticles in drug delivery, Journal of Controlled Release 100, 5-28, 2004. 20. J. Panyam, V. Labhasetwar, Biodegradable nanoparticles of drug and gene delivery to cells and tissue. Advanced Drug Delivery Reviews 55, 329-347, 2003. 21. G. Borchard, Chitosans for gene delivery, Advanced Drug Delivery Review 52, 145-150, 2001. 22. Hai-Quan mao, K.Roy, Kam.W. Leong, Chitosan-DNA nanoparticles as gene carriers: synthesis, characterization and transfection efficiency, Journal of Controlled release 70, 399-421, 2001. 23. S. Shirashi, T. Imai, M. Otagiri, Controlled release of indomethacin by chitosan-polyelectrolyte complex: optimization and in vivo: invitroevaluation Journal of Controlled release 25, 113-116, 1993. 24. Q.Gan, T.Wang, C.Cochrane, P. McCarron, Modulation of surface charge, particle size and morphological properties of chitosan-TPP nanoparticles intended for gene delivery, Colloids and surfaces B: Biointerfaces 44, 65-73, 2005. 25. D. Halliday, R. Resnick, J. Walker, Fundamentals of physics, John wiley and Sons, Inc. Canada, Fourth edition, 639-640, 1993. 26. H.S. Yoo, J.E. Lee et al., Self-assembled nanoparticles containing hydrophobically modified glycol chitosan for gene delivery, Journal of controlled release 103, 235-243, 2005. 27. S. Mansouri, P. Lavigne, K. Corsi, M. Benderdour, E. Beaumont, J.C. Fernandes, Chitosan-DNA nanoparticles as non-viral vectors in gene therapy: strategies to improve transfection efficacy, European Journal of Pharmaceutics and Biopharmaceutics 57, 1-8, 2004. 28. T. Ishii, Y. Okahata, T. Sato, Mechanism of cell transfection with plasmid/chitosan complexes, Biochimica. biophysica acta 1514, 51-64, 2001. 29. Yong Woo Cho, Jong-Duk Kim, Kinan Park, Polycation gene delivery system: escape from endosomes to cytosal, JPP 55, 721-734, 2003. 30. P. Midoux, M. Monsigny, Efficient Gene Transfer by Histidylated Polylysine-pDNA Complexes, Bioconjugate Chem 10, 406-411, 1999. 31. I.Fajac, P.Midoux, Histidylated polylysine as a synthetic vector for gene transfer into immortalized cystic fibrosis airway surface and airway gland serous cells, The journal of gene medicine 2, 368-378, 2000. 32. J.M. Benns, J.S. Choi, R.I. Mahato, J.S. Park, S.W. Kim, pH-sensitive cationic polymer gene delivery vehicle: N-Ac-poly(L-histidine)-graft-poly(L-lysine) comb shaped polymer, Bioconjugate Chem, 637-645, 2000. 33. E.Curotto, F.Aros, Quantitative-determination chitosan and the percentage of free amino-group, Anal. Biochem. 211, 240-241, 1993. 34. Leggett Bailey.J. in Techniques in Protein chemistry, Elsevier. Amsterdam, 348-349, 1967. 35. T. Sato, T. Ishii, Y. Okahata, In vitro gene delivery mediated by chitosan. Effect of pH, serum, and molecular mass of chitosan on the transfection efficiency, Biomaterial 22, 2075-2080, 2001. 36. C. R. Dass. Lipoplex-mediated delivery of nucleic acids: factors affecting in vivo transfection, J. Mol. Med. 82, 579-591, 2004. | |
| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/31658 | - |
| dc.description.abstract | 現在有許多蛋白質藥物量小且價格昂貴，生產的方式不外乎將所欲表現蛋白質的外源基因利用各種方式嵌入微生物（如E.coli、酵母菌等），但不能夠大量生產，且還有許多外源基因在微生物上的表現會受限、耗時費力等問題存在，所以，在基因轉殖動物的研究便成為人們注意的新焦點。
目前在基因轉殖雞的研究上，發現雞不但可以克服哺乳動物上會產生的缺點，而且還有許多額外的優勢在。 本研究在幾丁聚醣上接犬尿酸來當作一新的基因載體，並做出粒徑150~300 nm穩定均勻的奈米微粒，且DNA包覆率可達90 %以上。在N/P=2時，界面電位可達+9 mV，且比未改質幾丁聚醣更能有效保護DNA免於限制脢的切割。改質過後的幾丁聚醣對細胞並無負面影響，且能在3T3及HeLa細胞有轉染的表現，但3T3細胞轉染效果明顯優於HeLa，顯示基因載體對不同細胞的轉染能力不同。最後在雞胚於第0日時將此改質過後的基因載體注入雞胚，發現雞隻於奈米微粒注入後存活率剩下14%，且沒有綠色螢光表現，但改質後幾丁聚醣/DNA奈米微粒仍具有許多優勢可作為基因轉殖雞研究的載體。 | zh_TW |
| dc.description.abstract | There are many therapeutic proteins and peptide have been licensed for production using bacterial, fungal and mammalian cells, and many therapeutic proteins are currently being developed and tested in a variety of host cells. Transgenic animals describes animals with chromosomes that contain stably integrated copies of genes. The use of transgenics as a technology for producing recombinant proteins has made remarkable strides in the past few years. Recently, transgenic chickens should be near-perfect bioreactors for making large amounts of pure recombinant proteins.
In this study, we tried to use urocanic acid to modify chitosan as a new vector in gene delivery. We made the nanoparticles with diameter is 150~300 nm, and the DNA loading efficiency is over 90 %. When N/P=2, the zeta potential is +9 mV. And the modified chitosan/DNA nanoparticles can protect the DNA efficiently than unmodified chitosan/DNA. The modified chitosan had very good biocompatibility in 3T3 and HeLa, but the transfection efficiency is cell-dependant. In vivo test, the viability of the treated chicken embryos was down to 14%, and no any green fluroence protein was observed. But the modified chitosan/DNA nanoparticle still has a lot of advantages which applied in transgenic chicken. | en |
| dc.description.provenance | Made available in DSpace on 2021-06-13T03:16:53Z (GMT). No. of bitstreams: 1 ntu-95-R93548031-1.pdf: 84490559 bytes, checksum: 0c31736aa129c6ffc68d444131d3c1bd (MD5) Previous issue date: 2006 | en |
| dc.description.tableofcontents | 摘要 …………………………………………………………………………I
英文摘要 ……………………………………………………………………II 目錄 …………………………………………………………………………III 圖目錄 ………………………………………………………………………VI 表目錄 ………………………………………………………………………IX 第一章 前言 …………………………………………………………1 1-1 醫葯用蛋白的生產 …………………………………………………1 1-2 基因轉殖動物 ………………………………………………………1 1-3 各種將外源基因轉染至細胞的方法 ………………………………4 1-3-1 原核顯微注射法 ……………………………………………4 1-3-2 胚幹細胞載體法 ……………………………………………5 1-3-3 反轉錄病毒轉染法 …………………………………………6 1-3-4 精子載體法 …………………………………………………7 1-3-5 體細胞核轉置法 ……………………………………………8 1-4 基因轉殖雞 ………………………………………………………9 1-4-1 雞蛋與蛋白的成份 …………………………………………9 1-4-2 家禽基因轉殖的時機 …………………………………………10 1-4-3 基因轉殖雞的優勢 ……………………………………………13 1-4-4 基因轉殖雞的展望 ……………………………………………14 1-5 基因遞送系統 ……………………………………………………15 1-5-1 病毒載體及非病毒載體 ………………………………………15 1-5-2 幾丁聚醣 ………………………………………………………16 1-5-3 幾丁聚醣製配奈米微粒之方法 ………………………………17 1-5-3-1 乳化交聯法 ………………………………………………17 1-5-3-2 共生沉澱法 ………………………………………………18 1-5-3-3 噴霧乾燥法 ………………………………………………19 1-5-3-4 乳化液滴聯合法 …………………………………………20 1-5-3-5 離子凝膠法 ………………………………………………21 1-5-3-6 反微胞法 …………………………………………………22 1-5-3-7 篩選法 ……………………………………………………23 1-6 研究目的 ……………………………………………………………24 第二章 理論基礎 ……………………………………………………25 2-1 Coulomb’s Law ……………………………………………………25 2-2 基因載體進入細胞的機制 …………………………………………26 2-3 質子海綿效應 ………………………………………………………27 第三章 實驗方法……………………………………………………………30 3-1 實驗儀器 ……………………………………………………………30 3-2 實驗藥品 ……………………………………………………………31 3-3 實驗流程及材料製備 ………………………………………………32 3-3-1 犬尿酸接枝於幾丁聚醣 ………………………………………33 3-3-2 奈米微粒製備 …………………………………………………35 3-4 材料分析、生物可行性測試、體內、體外實驗 …………………37 3-4-1 FTIR分析 ………………………………………………………37 3-4-2 Ninhydrin 測試 ………………………………………………37 3-4-3 大量質體之萃取 ………………………………………………39 3-4-4 測量粒徑及界面電位（zeta potential )………………………39 3-4-5 穿透式電子顯微鏡（TEM）分析 ……………………………39 3-4-6 掃描式電子顯微鏡（SEM）分析 ……………………………39 3-4-7 DNA包覆效率測試 ……………………………………………40 3-4-8 電泳分析 ………………………………………………………41 3-4-9 WST-1細胞增生測試 …………………………………………41 3-4-10 細胞毒性測試 ………………………………………………43 3-4-11 體外轉染細胞實驗 …………………………………………44 3-4-12 動物實驗 ……………………………………………………45 第四章 結果與討論 …………………………………………………………47 4-1 幾丁聚醣接上犬尿酸 ………………………………………………47 4-1-1 FTIR分析結果…………………………………………………47 4-1-2 Ninhydrin 測試結果 …………………………………………49 4-2 幾丁聚醣奈米微粒製備 ……………………………………………51 4-2-1 粒徑分析 ………………………………………………………51 4-2-2 TEM、SEM分析 ………………………………………………56 4-2-3 界面電位 ………………………………………………………61 4-3 包覆效率測試 ………………………………………………………62 4-4 奈米微粒抗限制脢作用之電泳分析 ………………………………64 4-5 生物相容性測試 ……………………………………………………67 4-6 體外細胞轉染實驗 …………………………………………………72 4-7 動物實驗 ……………………………………………………………78 第五章 結論 ………………………………………………………………80 第六章 未來研究 ……………………………………………………………81 第七章 參考文獻 ……………………………………………………………82 | |
| dc.language.iso | zh-TW | |
| dc.subject | 幾丁聚醣 | zh_TW |
| dc.subject | 基因轉殖雞 | zh_TW |
| dc.subject | 奈米微粒 | zh_TW |
| dc.subject | 基因載體 | zh_TW |
| dc.subject | transgenic chicken | en |
| dc.subject | chiitosa | en |
| dc.subject | nanoparticle | en |
| dc.subject | gene delivery | en |
| dc.title | 幾丁聚醣接枝犬尿酸/DNA奈米微粒在基因轉殖雞上的應用 | zh_TW |
| dc.title | The application of chitosan-UA/DNA nanoparticles as gene carrier in transgenic chicken | en |
| dc.type | Thesis | |
| dc.date.schoolyear | 94-2 | |
| dc.description.degree | 碩士 | |
| dc.contributor.oralexamcommittee | 鄭登貴,郭宗甫,姚俊旭,張淑真 | |
| dc.subject.keyword | 幾丁聚醣,奈米微粒,基因載體,基因轉殖雞, | zh_TW |
| dc.subject.keyword | chiitosa,nanoparticle,gene delivery,transgenic chicken, | en |
| dc.relation.page | 85 | |
| dc.rights.note | 有償授權 | |
| dc.date.accepted | 2006-07-31 | |
| dc.contributor.author-college | 工學院 | zh_TW |
| dc.contributor.author-dept | 醫學工程學研究所 | zh_TW |
| 顯示於系所單位： | 醫學工程學研究所 | |
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