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Please use this identifier to cite or link to this item: http://hdl.handle.net/11455/52089
標題: 枯草桿菌xylose誘導型系統以及融合廣效性流感病毒HA抗原疫苗之枯草桿菌表現研究
Study on Bacillus subtilis xylose induce system and expression of fusion broad-spectrum influenza HA epitope vaccine by Bacillus subtilis
作者: Wu, Po-Yen
吳柏彥
關鍵字: HA高保留性抗原;HA highly-conserved antigen;融合蛋白;枯草桿菌表現系統;枯草桿菌雙質體系統;靈芝免疫調節蛋白;第一型重組蛋白類似物1A;fusion protein;Bacillus subtilis expression system;Bacillus subtilis dual plasmid system;Ganoderma lucidum immune regulatory protein LZ-8;recombinant type Ιantifreeze protein analogue
出版社: 食品暨應用生物科技學系所
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摘要: 
Bacillus subtilis is recognized as GRAS (generally recognized as safe) bacteria and has been applied in many industries, such as fermented food , enzymes, and special chemicals. Bacillus subtilis has been used as a probiotic or as plant disease control agents. At 2009, a global outbreak of a new strain H1N1 influenza virus was spreed in just a few months. The global pandemic quickly lead to serious disaster. The main reason is that the traditional manufacture of influenza virus vaccine only against the targeted virus strain, and preparation of enough vaccines take several months. A broad-spectrum influenza vaccine thus been considered as a new strategy. In this study, the highly conserved influenza virus hemagglutinin HA2 N terminal 14 amino acid fragment was choosen as antigen, and the gene was synthesized by PCR. The epitope was fused to Previously constructed Ganoderma lucidum immune regulatory protein LZ-8 as carrier protein and the recombinant antifreeze protein analogue 1A as linker, the designed LZ8-1A-HA(e) recombinant protein was expressed by Bacillus subtilis expression system. The LZ8-1A-HA(e) was used to immunize mice to examine the potency as influenza vaccine.
Another part of this study, aims to decrease the basal level of previously conststructed Bacillus subtilis xylose inducible dual plasmid system. However, in the 3 constructed system, the basal level did not decrease as expected.

枯草桿菌(Bacillus subtilis)為GRAS (generally recognized as safe)級菌種，已應用在許多產業上，如在發酵食品之製造、酵素及特用化學品之生產、作為益生菌（probiotic）及作為植物病害防治試劑等。2009年所爆發的H1N1新型流感病毒，在短短數個月的時間於全球快速傳播，造成大流行導致嚴重災情。由於傳統流感病毒疫苗只能針對特定製造疫苗病毒株，而且疫苗製備需耗時數月。發展廣效性流感疫苗成為現今新的治療流感病毒策略。本實驗設計以流感病毒中高度保留的血球凝集素HA2 N端14個胺基酸抗原片段，並融合了本實驗室之前所構築的重組靈芝免疫調節蛋白LZ-8與第一型重組抗凍蛋白類似物1A，設計出LZ8-1A-HA(e) 重組融合蛋白，利用枯草桿菌表現系統生產，並評估以枯草桿菌表現系統生產出的LZ8-1A-HA(e) 作為疫苗之效果，進行動物試驗檢測其抗體產生情形。結果顯示LZ8-1A-HA(e) 可誘發針對LZ8、1A與LZ8-1A-HA(e) 與2009年大流行性流感病毒疫苗(A/H1N1) HA抗原的特異性抗體。
另一方面，在之前本實驗室所構築的枯草桿菌xylose雙質體誘導系統上，存在著基礎表現量過高的問題，所以本實驗另一部分試圖改善枯草桿菌xylose誘導型雙質體系統於未添加入誘導物的基礎表現量，結果在搭配不同表現質體與調控質體的菌株其有無添加誘導物的表現量毫無差異。
URI: http://hdl.handle.net/11455/52089
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