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研究生:Urantsetseg Gantulga
研究生(外文):Urantsetseg Gantulga
論文名稱:建立表現蟑螂重組Per a 3 抗原蛋白之轉基因菸草細胞培養
論文名稱(外文):Establishment of Cell Culture System of Transgenic Tobacco Expressing the Cockroach Recombination Per a 3 allergen
指導教授:余聰安
指導教授(外文):Tsong Ann Yu
口試委員:李泰林江主惠余聰安
口試日期:2018-07-19
學位類別:碩士
校院名稱:大葉大學
系所名稱:分子生物科技學系碩士班
學門:生命科學學門
學類:生物化學學類
論文種類:學術論文
論文出版年:2018
畢業學年度:106
語文別:英文
論文頁數:25
中文關鍵詞:轉基因菸草Per a 3癒傷組織細胞培養
外文關鍵詞:Transgenic tobaccoCockroach the Per a 3 geneCallus inductionCell culture system
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蟑螂是造成全世界哮喘的重要原因,德國蟑螂(Blattella germanica)和美洲蟑螂(Periplaneta Americana)是最具代表性蟑螂物種。Per a 3是美國蟑螂的重要過敏原之一,利用轉基因植物生產Per a 3,具有成本低,易於擴展生產和安全的優點。本研究目的是將已轉Per a 3基因煙草,經由細胞培養的模式,建立一套有效率生產Per a 3的系統,希望對後續Per a 3蛋白的生產有有所助益。轉Per a 3基因菸草植物,移植到MS培養基,培養在24±1℃,70%-80%,30 μmol/m2/s的條件下,每隔4週繼代培養一次,做為日後的培養材料。將葉片培植體培養在含有不同2.4-D和BA濃度的MS培養基,以誘導癒傷組織,每隔4週繼代,結果顯示在含有0.2 mg / l 2.4-D和0.5 mg / l BA培養基,最適合癒傷組織的誘導。將這些癒傷組織轉移到含同樣成分的液體培養基培養,並每週進行細胞培養的評估,結果顯示,細胞組織懸浮體積每週增加2 ml。
Cockroach is a significant cause of asthma worldwide, with the most relevant cockroach species is the German cockroach (Blattella germanica) and American cockroach (Periplaneta Americana). Per a 3 is one of the important allergens of the American cockroach (Periplaneta americana). Using a transgenic plant aprroach produing recombination Per a 3 has some advantages such as low cost, easily scalable production, and safety. The objective of this study was developed an effective system for the cell culture of transgenic tobacco expressing Per a 3 gene. Previously, the transgenic tobacco with Per a 3 gene has been generated in our laboratory. Transgenic tobacco plants in vitro were culture under the conditions of 24 ± 1˚C , 70% - 80% RH, and light intensity of 30 µmol/m2/. s. The leaves explants were cut and cultured on a solid Murashige & Skoog’s (MS) medium containing the different concentration combinations of 2,4-D Dichlorophenoxyacetic acid (2,4-D) and 6-Benzylaminopurina (BA) for callus induction, and sub-cultured very four weeks. The data showed the best suitable medium for callus was the MS medium adding 0.2 mg/l 2,4-D and 0.5 mg/l BA. The calli were transferred into the same liquid MS medium for cell suspension cultures and detected the cell gowth rates very one week. Our result indicated that the cell suspension volume were increased two ml every week.
封面內頁
簽名頁
中文摘要 iii
ABSTRACT iv
ACKNOWLEDGMENTS v
TABLE OF CONTENTS vi
LIST OF FIGURES viii
LIST OF TABLES ix
LIST OF ABBREVIATIONS x

CHAPTER 1. INTRODUCTION 1
1.1 American cockroach allergen 1
1.1.1 Cockroach the Per a 3 allergen 2
1.1.2 Recombinant Allergen Protein expression in Plants cell Culture 3
1.2 Per a 3 protein expression in transgenic tobacco callus 4
1.3 The cell culture using expressing the target gene 6
CHAPTER 2. MATERIALS AND METHODS 8
2.1. Experimental materials 8
2.1.1. Antibiotics 8
2.1.2. Hormones 8
2.1.3. Murashige & Skoog’s medium 9
2.2. Establishment of cell culture for the transgenic tobacco cell 9
2.3. The effects of different plant regulars on a transgenic tobacco callus 10
CHAPTER 3. RESULTS 11
3.1. Establishment of cell culture for the transgenic tobacco cell 11
3.2. The effects of different plant regulars on a transgenic tobacco callus 11
CHAPTER 4. DISCUSSION AND CONCLUSIONS 13
REFERENCES 22


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