轉錄體分析 Acetobacterium woodii 降解1,1,1-三氯乙烷機制並用以協助 Dehalococcoides進行還原脫氯

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林志豪(Chih-Hao Lin) 查詢紙本館藏

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轉錄體分析 Acetobacterium woodii 降解1,1,1-三氯乙烷機制並用以協助 Dehalococcoides進行還原脫氯
(The degradation mechanisms of Acetobacterium woodii toward 1,1,1-trichloroethane by transcriptome analysis and its role in assisting reductive dechlorination of Dehalococcoides)

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摘要(中)

1,1,1-三氯乙烷(1,1,1-Trichloroethane)是地下水污染的常見來源之一，不僅導致人類患癌，也抑制Dehalococcoides mccartyi(D. mccartyi) 進行三氯乙烯(Trichloroethene)的還原脫氯反應，而三氯乙烯也是主要地下水污染物之一。在研究中Acetobacterium woodii (A. woodii) 具有1,1,1-三氯乙烷的降解能力。然而，目前對於其降解機制仍不清楚。在本研究中欲探討在果糖生長下的A. woodii降解1,1,1-三氯乙烷時所參與的代謝途徑以及嘗試透過在1,1,1-三氯乙烷和三氯乙烯共汙染下共培養A. woodii和D. mccartyi以消除1,1,1-三氯乙烷抑制三氯乙烯脫氯作用。結果顯示，A. woodii能夠降解最高至2 mM的 1,1,1-三氯乙烷。為了進一步探討其潛在機制，我們使用轉錄組分析和RT-qPCR方法。結果表明，1,1,1-三氯乙烷誘導了22個與Wood-Ljungdahl途徑相關基因的上調表達，推測其可能與1,1,1-三氯乙烷的降解有相關性。為了減少1,1,1-三氯乙烷對D. mccartyi進行三氯乙烯脫氯的負面影響，接著我們建立了一個由D. mccartyi CWV2和A. woodii組成的共培養系統，該系統能夠協同降解1,1,1-三氯乙烷和三氯乙烯。本研究不僅為探討了A. woodii對1,1,1-三氯乙烷的降解機制提供了新的見解，也在1,1,1-三氯乙烷和三氯乙烯共污染的場地中利用A. woodii進行生物修復提供了多種策略。

摘要(英)

1,1,1-Trichloroethane (1,1,1-TCA), one of the common sources of groundwater contamination, not only causes cancer in humans but also inhibits the reductive dechlorination of trichloroethene (TCE) by Dehalococcoides mccartyi (D. mccartyi), which is the major groundwater contamination. Previous studies have reported that Acetobacterium woodii (A. woodii) exhibits 1,1,1-TCA degradation activity. However, the degradation mechanism remains unclear. This study aimed to identify for the key pathways involved in the degradation of 1,1,1-TCA by A. woodii when grown on fructose as a primary substrate, and to establish a co-culture consisting of D. mccartyi and A. woodii to alleviate the inhibitory effect of 1,1,1-TCA on the dechlorination of TCE. The results showed that A. woodii can degrade 2 mM of 1,1,1-TCA. To investigate the underlying mechanisms of 1,1,1-TCA degradation, we adopted transcriptomic analysis and RT-qPCR methods. The results indicated that 1,1,1-TCA induced the up-regulation of twenty-two genes related to Wood-Ljungdahl pathway, suggesting that this pathway was involved. To alleviate the negative impact of 1,1,1-TCA on TCE degradation by D. mccartyi, we established a co-culture consisting of D. mccartyi strain CWV2 and A. woodii to synergistically degrade both 1,1,1-TCA and TCE by providing fructose as the sole carbon source. This study not only provides new insight to reveal the degradation mechanism of 1,1,1-TCA by A. woodii, but also provide strategies for removing 1,1,1-TCA by A. woodii and then dechlorination of TCE in the both 1,1,1-TCA and TCE co-contaminated sites.

關鍵字(中)

★ Acetobacterium woodii
★ 1,1,1-三氯乙烷
★ 共培養
★ Dehalococcoides
★ 共汙染

關鍵字(英)

★ Acetobacterium woodii
★ 1,1,1-TCA
★ co-culture
★ Dehalococcoides
★ co-contamination

論文目次

國立中央大學圖書館學位論文授權書 i
國家圖書館學位論文延後公開申請書 ii
國立中央大學碩士班研究生論文指導教授推薦書 iii
國立中央大學碩士班研究生論文口試委員審定書 iv
致謝 v
摘要 vi
Abstract vii
目錄 viii
圖目錄 x
表目錄 xii
第壹章緒論 (Introduction) 1
1.1 研究動機 1
1.2 1,1,1-三氯乙烷抑制多氯乙烯還原脫氯 2
1.3 1,1,1-三氯乙烷之厭氧生物降解 2
1.4 Acetobacterium 降解有機鹵化物 3
第貳章實驗目的及架構 6
第参章實驗材料與方法 (Materials and Methods) 8
3.1 實驗材料 8
3.1.1 常用藥品與試劑 8
3.2 實驗方法 11
3.2.1 DCB-1 厭氧菌培養基配置 11
3.2.2 Acetobacterium spp.菌株的保存與來源 11
3.2.3 Dehalococcoides mccartyi菌株的保存與來源 13
3.2.4 細菌Genomic DNA 萃取 14
3.2.5 氣相層析儀分析 15
3.2.6 定量聚合酶連鎖反應 (qPCR) 16
3.2.7 細菌RNA萃取 16
3.2.8 RNA反轉錄 17
3.2.9 醋酸濃度測定 18
3.2.10 轉錄組分析 19
第肆章實驗結果 (Result) 21
4.1 Acetobacterium woodii共代謝降解1,1,1-三氯乙烷 21
4.2 轉錄體分析Acetobacterium降解時1,1,1-三氯乙烷下基因表現 22
4.3 Dehalococcoides mccartyi與Acetobacterium woodii共培養協同降解1,1,1-三氯乙烷及三氯乙烯 23
第伍章討論(Discussion) 26
5.1 Acetobacterium woodii降解1,1,1-三氯乙烷時之生理特性 26
5.2 分析Acetobacterium woodii潛在參與降解1,1,1-三氯乙烷之代謝路徑 27
5.3 Dehalococcoides 藉由與Acetobacterium woodii 共培養消除1,1,1-三氯乙烷抑制三氯乙烯脫氯反應之效果 30
第陸章結論 (Conclusion) 32
參考文獻 (Reference) 33
圖表 39
附加資料 (Supplementary Data) 54

參考文獻

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指導教授

陳師慶

審核日期

2023-7-20

推文