以變性梯度膠體電泳法分析土壤中酚降解菌之變化

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Title:	以變性梯度膠體電泳法分析土壤中酚降解菌之變化 Analysis of the Changes of Phenol Degraders by Denaturing Gradient Gel Electrophoresis During Phenol Degradation in soils
Authors:	楊如玉 Ju-Yu Yang
Contributors:	荊樹人劉瑞美嘉南藥理科技大學：環境工程與科學系碩士班
Keywords:	變性梯度膠體電泳酚降解菌 Phenol Degraders Denaturing Gradient Gel Electrophoresis
Date:	2005
Issue Date:	2008-10-31 16:15:17 (UTC+8)
Abstract:	酚在工業上是常見的污染之一，其苯環結構穩定需要外力或是微生物作用，才能將其開環，成為長鏈之結構以便後續分解程序，生物降解方式因為較具經濟上的優勢，故可選用做為土壤酚污染整治最佳選擇之一。本實驗自欣岱股份有限公司的反應釜及廢水儲槽周圍收集污泥或土壤，以批次培養方式，篩選出耐高濃度之酚降解菌M1、M5，並對篩選出之純菌菌株進行革蘭氏染色，及鄰苯二酚酵素測定。結果顯示M1、M5菌株皆為革蘭氏陽性菌，具有鄰苯二酚1,2-二加氧酶活性，經過16S rDNA定序比對，M1為Ralstonia basilensis(AB109778)相似度96%，M5為Staphylococcus sp. B60(AF333342)相似度94%。另外將實驗菌株接種於模擬污染土壤中，進行批次生物降解實驗，監測實驗過程酚濃度及酚降解菌之菌數，同時進行土壤DNA萃取，經引子341GC+534r作PCR反應，以變性梯度膠體電泳分析，藉此追蹤實驗過程中菌株之消長變化。實驗結果發現接種試驗菌株於模擬污染土壤中均具良好降解效果，且於台糖研究所土壤中有較佳的降解效果。應用變性梯度膠體電泳分析，菌株之指紋圖譜亮帶隨著降解時間增加而增加，間接表示菌落數的增加，顯示變性梯度膠體電泳分析比傳統培養方式更具敏感度。 Phenol is a common pollutant from industries, it impacts on the aquatic ecosystem and environmental balance extremely, and biodegradation is a very economic technique, which is the best choice of the phenol contaminated soil remediation. The phenol high-tolerance degraders were isolated from collected sludge and soil from petrol station and around of reactor and wastewater storage in Syndyne Industry, the tested strains in this study including M1 and M5. The physiological properties of te sted strains were analyzed by Gram stain, activity of catechol dioxygenase. The results showed that the tested strains were Gram positive and had the activities of catechol 1,2-dioxygenase . The M1 and M5 were showed 96% probability of Ralstonia basilensis (AB109778) and Staphylococcus sp. B60 (AF333342) analysed in the denaturing gradient gel electrophoresis (DGGE) profiles of amplified 16S rDNA fragment. Inoculation of effective degraders to the modified polluted soils spiked with phenol was carried out by batch culture. The phenol concentration and the number of phenol degrader were monitored in the experimental period, soil DNA was extracted and amplified by primer 341GC+534r of PCR for DGGE analysis at the same time, which to understand the variation of test strains. The results showed that the biodegradation in the polluted soils and aquatic m-cresol solution were not consistent. All the strains can degrade phenol in the test polluted soils efficiently, the most effective degradation in the soil collected from was observed by test strains. Tracing soil bacterial community by DGGE technology in the experimental period was observed that inoculated effective strains became dominant, and this result was similar to the colony forming unit (CFU) was counted by plate-count methods. The denaturing gradient gel electrophoresis (DGGE) is considered one of the most sensitive of the scanning techniques for soil community.
Relation:	校內外完全公開
Appears in Collections:	[Dept. of Environmental Engineering and Science (including master's program)] Dissertations and Theses

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