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    請使用永久網址來引用或連結此文件: http://ir.lib.ncu.edu.tw/handle/987654321/72877


    題名: Pseudomonas sp. A46全基因組分析與重金屬復育基因工程菌開發;Genomic Characterization of Pseudomonas sp. A46 and Establishment of Genetic Bacteria for Enhancing Bioremediation of Heavy Metals
    作者: 羅凱泓;Luo, Kai-Hong
    貢獻者: 生命科學系
    關鍵詞: ;重金屬;生物復育;次世代定序;金屬硫蛋白;基因工程菌;mercury;heavy metal;bioremediation;next generation sequencing;metallothionein;genetic bacteria
    日期: 2017-01-06
    上傳時間: 2017-05-05 17:12:54 (UTC+8)
    出版者: 國立中央大學
    摘要: 汞是一種自然存在的元素,常溫下唯一的液態金屬,帶有強烈的生物毒性。環境中有許多細菌具有汞抗性,能抵抗汞汙染環境並將高毒性型態汞轉化成低毒性型態。抗汞菌株Pseudomonas sp. A46能培養於含60 ppm Hg2+ LB培養基中,並於24小時內移除培養基中89.24%中汞離子,推測可為汞汙染生物復育之優勢菌株。本論文利用次世代定序技術分析Pseudomonas sp. A46基因體組成,分析結果顯示除mer operon外可針對汞進行生物復育外,菌株還帶有鎘抗性基因czcABC、鉻抗性與還原基因chr operon、銅抗性和氧化基因copB與cueO、鉛抗性基因ZntA與砷抗性和還原基因acr3與arsC,因此此菌株除汞外可能帶有上述重金屬之抗性與生物復育潛力可供外來進一步研究。有研究指出重金屬會對細胞造成氧化壓力,qPCR分析結果顯示菌株培養在含汞環境下,除汞操作組外其抗氧化相關基因如SOD、GPx、CAT等表現量皆有上升。由於利用汞操作組還原揮發汞,經過汞循環路徑,最終仍可能於其它地區沉降,無法完全解決汞汙染問題,本論文參考前人文獻在Pseudomonas sp. A46菌株中表現能結合重金屬之金屬硫蛋白,將重金屬固定於菌株內以便後續回收,此外為解決蛋白質表現問題,本論文開發Pseudomonas sp. A46用pJBR持續表現型蛋白質表現系統,並將綠螢光蛋白與金屬硫蛋白融合,使金屬硫蛋白能於菌株中穩定表現並能簡易觀察蛋白質表現量,汞固定實驗顯示轉殖金屬硫蛋白之Pseudomonas sp. A46菌體內汞濃度較野生型高3.7倍,顯示利用金屬硫蛋白增加汞吸收量是可行的。;Mercury is a nature liquid element which has strong toxicity. In the environment, many mercury-resistant bacteria can transform the high-toxic state mercury into low-toxic state mercury. One of the mercury-resistant bacteria, Pseudomonas sp. A46, is able to growth in the LB medium with 60ppm mercuric ion, to remove 89.24% mercuric ion from the medium in 24 hours. We suggested that P. spp A46 can be the potential strain for mercury-bioremediation. Using the Next Generation Sequencing (NGS) technique to analyze whole genome of P. spp A46, our results indicated that the strain not only carries mer operon for mercury-bioremediation but also cadmium resistant gene czcABC、chromium resistant and reduction gene chr operon、copper resistant and oxidation gene copB and cueO、lead resistant gene zntA and arsenic resistant and reduction gene acr3 and arsC. As mercury was reported to induce the oxidative stresss, our results also confirmed that the expression levels of superoxide dismutase, glutathione peroxidase and catalase were increased under mercury stress. Although mercury was potentially removed by Pseudomonas sp. A46, part of mercury could be vaporized into air. To avoid the vaporization of mercury when bioremediation was operating, we developed one genetic bacteria harbouring metallothionein, a heavy metal-binding protein. Our results found out that the gentic bacteria, P. spp A46, could bind to mercury with 3.7 fold concentration of mercury than wild type strain.
    顯示於類別:[生命科學研究所 ] 博碩士論文

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