Please use this identifier to cite or link to this item: http://hdl.handle.net/11455/92361
標題: Chlorella sp. DT蝦紅素生產之培養條件最適化並利用轉基因菸草生產蝦紅素
Optimize the cultural condition for astaxanthin production in Chlorella. sp. DT and produce astaxanthin by transgenic plant
作者: Yi-Ting Kuo
郭一廷
關鍵字: Chlorella sp. DT
Astaxanthin
β-carotene ketolase
Chlorella sp. DT
蝦紅素
β-carotene ketolase
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摘要: The Chlorella sp. DT, an endemic species in Taiwan, was found with astaxanthin producing ability. To maximize productivity of astaxanthin in Chlorella sp. DT, NaNO3, Na2CO3 and CaCl2 were selected as inducers by factors screening. Further, response surface methodology was used to optimize the cultural condition for astaxanthin production. The results showed astaxanthin productivity was 0.749 mg L-1 after 10 days of cultivation and induction by used the optimized the cultural condition, BG11 medium with NaNO3 2779.08 mg L-1, Na2CO3 273.97 mg L-1 and CaCl2 75.34 mg L-1. Furthermore, based on factors screening, the culture conditions for two-stage to produce astaxanthin by Chlorella sp. DT were developed. The PBD-2 medium was used to accumulate biomass in green-stage, and the Pred-medium was used for astaxanthin production in induce-stage. Astaxanthin productivity is 0.805 mg L-1 after 12 days of cultivation and induction in two-stage strategy. We successfully maximize the astaxanthin production in Chlorella sp. DT by one and two-stage of cultivation strategies. To increase the production of astaxanthin, we cloned the Chlorella sp. DT β-carotene ketolase (BKT), the key enzyme of astaxanthin synthesis, and overexpressed the BKT gene in Nicotiana tabacum and N. benthamiana by using agrobacterium-mediated transformation. The transgenic N. tabacum and N. benthamiana plants can produce 604.6 and 385.8 μg astaxanthin per g (dry weightof leaves), respectively. This study successfully used β-carotene ketolase gene form Chlorella sp. DT to build a transgenic plant system for astaxanthin production.
台灣原生的藻種Chlorella sp. DT具有形成蝦紅素的能力。為了進一步探討Chlorella sp. DT最大蝦紅素產量,本研究利用實驗設計法篩選出有助增加藻體中蝦紅素含量的培養因子:硝酸鈉、碳酸鈉及氯化鈣,更進一步以反應曲面法作最適化生產蝦紅素的培養基探討,結果顯示添加2779.08 mg L-1的硝酸鈉、273.97 mg L-1的碳酸鈉以及75.34 mg L-1的氯化鈣於BG-11培養基,經過10天培養與誘導後蝦紅素產量為0.749 mg L-1。並由因子篩選探討的實驗結果,設計兩階段培養Chlorella sp. DT生產蝦紅素的培養基條件,以適合藻體生長的PBD-2培養基累積藻體量,再以Pred-medium培養基進行誘導蝦紅素生成,經過12天的培養與誘導後蝦紅素產量達0.805 mg L-1,實驗結果成功地以一階段及兩階段的培養方式探討Chlorella sp. DT最大的蝦紅素產量。並且為了提升蝦紅素產量,本研究也選殖了Chlorella sp. DT生成蝦紅素的關鍵蛋白質β-carotene ketolase,以農桿菌介導轉殖法將β-carotene ketolase表達至不具蝦紅素形成能力的Nicotiana tabacum及N. benthamiana中,大量表現β-carotene ketolase催化β類胡蘿蔔素代謝形成蝦紅素,使N. tabacum的T0轉殖株葉片組織生成604.6 μg g-1 DW的蝦紅素含量,N. benthamiana的T0轉殖株葉片組織生成385.8 μg g-1 DW的蝦紅素含量,成功以Chlorella sp. DT的β-carotene ketolase基因建立轉基因作物生產蝦紅素的平台。
URI: http://hdl.handle.net/11455/92361
其他識別: U0005-0302201514000400
文章公開時間: 10000-01-01
Appears in Collections:基因體暨生物資訊學研究所

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