啤酒去氫酶 ALD2p 和 ALD3p 為 ACDH (acetaldehyde dehyrogenase) 家族中的成員，並利用 NAD+ 作為輔酶。文獻指出，在泛酸合成的途徑中， ALD2p 和 ALD3p 會將中間產物 3-aminopropanal 轉變成為 β-alanine。

在哺乳類中， ALDHs 存在於角膜蛋白質中，以保護眼睛，透過直接吸收紫外光、代謝脂質過氧化物、抗氧化的功能保護眼睛。

由於先前實驗測試不到 ALD2p 及 ALD3p 的活性，所以在此測試是否有其他的功能，例如上述的抗紫外光、代謝脂質過氧化物、抗氧化的功能。
   
透過剔除 ALD2、ALD3 基因及過度表現 ALD2、ALD3 基因，接著再用滅菌過的超純水清洗，稀釋成不同濃度將菌液沾點於固態培養基上，照射紫外光或稀釋成不同濃度沾點於含有 H2O2 的固態培養基上。

In Saccharomyces cerevisia ALD2 (YMR170c) and ALD3 (YMR169c) are the members of ACDH (acetaldydehyde dehydrogenase) family, and the gene products are supposed to use NAD+ as coenzyme. Recent reference also shows that ALD2 and ALD3 genes are require for pantothenic acid biosynthesis by conversion of 3-aminopropanal to β-alanine.
	
In mammals, ALDHs have been identified as corneal crystallins and to contribute to eye protection by directly absorbing ultraviolet, or  detoxifying reactive aldehydes produced by lipid peroxidation to non-toxic metabolities and act as antioxidant.

In this study, we examined the effects of ultraviolet and peroxide by deletion and overexpression of ALD2 and ALD3 gene.

目錄
謝誌	I
中文摘要	II
英文摘要	IV
目錄	V
表目錄	VII
圖目錄	VIII
第一章 序論	1
第二章 實驗方法	5
2.1 儀器與設備 	5
2.2 藥品	7
2.3 實驗步驟	13
2.3.1誘導基因表現	13
2.3.2 超音波破碎細胞	14
2.3.3 SDS-PAGE 膠體電泳蛋白質分析法	14
2.3.4 膠體配製	15
2.3.5 蛋白質電泳操作	15
2.3.6 沾點顯現法 (點盤) 	17
2.3.6.1 紫外光對剔除 ALD2 及 ALD3 基因的酵母菌的影響	17
2.3.6.2 紫外光對過度表現 ALD2 及 ALD3 基因的酵母菌的影響	18
2.3.6.3 過氧化物對剔除 ALD2 及 ALD3 基因的酵母菌的影響	20
2.3.6.4 過氧化物對過度表現 ALD2 及 ALD3 基因的酵母菌的影響	21
2.3.6.5 過氧化物對剔除 ALD2 及 ALD3 基因的酵母菌的影響	23
2.3.6.6 過氧化物對過度表現 ALD2 及 ALD3 基因的酵母菌的影響	24
2.3.6.7 丙烯醛對剔除 ALD2 基因的酵母菌的影響	26
2.3.6.8 丙烯醛對過度表現 ALD2 基因的酵母菌的影響	27
2.3.6.9 乙醇對剔除 ALD3 基因的酵母菌的影響	29
第三章 結果與討論	30
第四章 結論	68
參考文獻	69
表目錄
表2.2.1 : YPD medium 組成	9
表2.2.2 : YPD plate 組成	9
表2.2.3 : YPD plate (各別含1 mM、 3 mM、 5 mM H2O2) 組成	9
表2.2.4 : SC-Ura (1% glucose) medium組成	10
表2.2.5 : SC-Ura (1%glucose) plate 組成	10
表2.2.6 : SC-Ura (1%raffinose) medium組成	10 
表2.2.7 : Separating gel 組成	11
表2.2.8 : Stacking gel 組成	11
表2.2.9 : Breaking buffer 組成	11
表2.2.10 : 2X staining buffer 組成	11
表2.2.11 : 1X Tris-glycine 電泳緩衝液組成	12
表2.2.12 : Coomassie Brilliant Blue R-250 組成	12
表2.2.13 : Destaining solution 組成	12
圖目錄
Fig. 1 PDH bypass 代謝途徑	1
Fig. 1.2 ALDHs 之功能	2
Fig. 1.3 合成泛酸的途徑及其過程中各基因的功能	3
Fig. 1.4 ALDH3A1 與 ALDH1A1 在角膜中的功能	4
Fig. 3.1 紫外光對剔除基因的影響，能量 3 × 103 μJ/cm2 (YPD 培養液清洗)	37
Fig. 3.2 紫外光對剔除基因的影響，能量 4 × 103 μJ/cm2 (YPD 培養液清洗)	38
Fig. 3.3 紫外光對剔除基因的影響，能量 5 × 103 μJ/cm2 (YPD 培養液清洗)	39
Fig. 3.4 紫外光對過度表現基因的影響，能量 3 × 103 μJ/cm2 (YPD 培養液清洗)	40
Fig. 3.5 紫外光對過度表現基因的影響，能量 4 × 103 μJ/cm2 (YPD 培養液清洗)	41
Fig. 3.6 紫外光對過度表現基因的影響，能量 5 × 103 μJ/cm2 (YPD 培養液清洗)	42
Fig. 3.7 紫外光對剔除基因的影響，能量 3 × 103 μJ/cm2 (滅菌過的超純水清洗)	43
Fig. 3.8 紫外光對剔除基因的影響，能量 4 × 103 μJ/cm2 (滅菌過的超純水清洗)	44
Fig. 3.9 紫外光對剔除基因的影響，能量 5 × 103 μJ/cm2 (滅菌過的超純水清洗)	45
Fig. 3.10 紫外光對過度表現基因的影響，能量 3 × 103 μJ/cm2 (滅菌過的超純水清洗)	46
Fig. 3.11 紫外光對剔除基因的影響，能量 4 × 103 μJ/cm2 (滅菌過的超純水清洗)	47
Fig. 3.12 紫外光對剔除基因的影響，能量 5 × 103 μJ/cm2 (滅菌過的超純水清洗)	48
Fig. 3.13 1 mM H2O2 對剔除基因的影響	49
Fig. 3.14 3 mM H2O2 對剔除基因的影響	50
Fig. 3.15 5 mM H2O2 對剔除基因的影響	51
Fig. 3.16 1 mM H2O2 對過度表現基因的影響	52
Fig. 3.17 3 mM H2O2 對過度表現基因的影響	53
Fig. 3.18 5 mM H2O2 對過度表現基因的影響	54
Fig. 3.19 1 mM H2O2 對剔除基因的影響	55
Fig. 3.20 3 mM H2O2 對剔除基因的影響	56
Fig. 3.21 5 mM H2O2 對剔除基因的影響	57
Fig. 3.22 1 mM H2O2 對過度表現基因的影響	58
Fig. 3.23 3 mM H2O2 對過度表現基因的影響	59
Fig. 3.24 5 mM H2O2 對過度表現基因的影響	60
Fig. 3.25 1 mM acrolein 對剔除及過度表現基因的影響	61
Fig. 3.26 3 mM acrolein 對剔除及過度表現基因的影響	62
Fig. 3.27 5 mM acrolein 對剔除及過度表現基因的影響	63
Fig. 3.28 6% ethanol 對剔除基因的影響	64
Fig. 3.29 8% ethanol對剔除基因的影響	65
Fig. 3.30 10% ethanol 對剔除基因的影響	66
Fig. 3.31 ALD2-pYes2 和 ALD3-pYes2 誘導及未誘導比較	67

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