臺灣博碩士論文加值系統

摘要

　　台灣西北部油氣田產油氣層絕大部份於中新世地層內發現，其中以中新世打鹿頁岩所夾儲油氣砂岩的產量最為豐富。地層中的微生物對於自然界物質的轉化和循環有著極為重要的影響。本研究自苗栗地區打鹿頁岩油氣井表土、地表下3200 m深的含油泥漿及3386 m深的岩心共分離出18株菌株，經TEM(穿透式電子顯微鏡)及革蘭氏染色得知所分離出之微生物型態主要為桿菌與短桿菌，而且菌落外觀也不盡相同，其中包含十四株革蘭氏陰性菌及四株革蘭氏陽性菌。利用16S rDNA基因序列的分析與參考菌株，比較菌種的親緣關係，並輔以API微生物鑑定套組測試生化特性鑑定菌種。在三個地層中皆有發現格蘭氏陰性菌的存在但型態皆不相同。此外，本研究亦探討溫度、pH、鹽度對菌株的影響。溫度方面F21、F30、TG12、TG31、SN10等菌株分離株，在溫度90℃皆還能存活，其中TG31在溫度90℃時還能夠有很好的繁殖能力；pH值方面在測試酸耐受度F21、F22、F31、TG20、TG31、TG32等菌株分離株，在<pH4時皆還能生長；鹼度方面僅TG11、TG12、FE20等菌株分離株，鹼耐受度為pH11，其餘鹼耐受度皆> pH11還能生長；鹽度方面以硫酸還原培養基培養之SR10、SR21、SR22、SR30等菌株分離株的耐受度以大於5%為最高。

Abstract
The deep oil-bearing Talu sandstone of the Miocene age was investigated for microbiological analyses in this study. A total of 18 bacterial strains were isolated from the surface soil and the downcore of 3380 m-deep Talu sandstone and 3200 m-deep drilling muds in an oil well of the Miaol area. When examined by transmission electron microscopey (TEM) and Gram’s stain, most of the bacterial isolates appeared to be rod-shaped or short rod-shaped. They also varied in the colonial morphology. The isolated strains were identified based on the bacterial phylogeny and the 16S rDNA gene sequence alignment with the reference strains in association with the biochemical assay using the API microbiology Kit. In the different depths (3200m and 3386m) of the downcore, all Gram-negative bacteria are not the same in morphology. The effects of the temperature, pH, and salinity on the growth of bacterial isolates were investigated. As for temperature, the FM21, FM30, TG12, TG31 and SN10 isolates could survive up to 90℃, in which TG31 could also be well reproducible at 90℃. As for pH, the FM21, FM22, FM31, TG20, TG31, and TG32 isolates could survive at <pH4, whereas the TG11, TG12, and FE20 isolates could survive up to pH11, and others could survive at >pH11. As for salinity, the SR10, SR21, SR22, and SR30 isolates could survive up to 50‰.

目 錄
摘要......................................................I
Abstract..................................................II
誌謝.....................................................III
目錄.....................................................IV
表目錄...................................................VI
圖目錄.................................................VIII
壹、緒論...................................................1
貳、文獻回顧...............................................3
2.1 石油有機成因學說....................................3
2.2 地質環境及沉積環境概況..............................5
2.3 細菌生態的研究......................................9
叄、實驗材料與方法........................................11
3.1 樣品來源...........................................11
3.2 培養基及相關試劑...................................11
3.3 器材...............................................13
3.4 儀器...............................................13
3.5 廠商細目...........................................14
3.6 菌種脫附...........................................15
3.7 培養基溶液的製備...................................15
3.8 DNA洋菜膠體電泳...................................20
3.9 聚合酶連鎖反應.....................................20
3.10 PCR產物DNA片段之回收與序列分析.................21
3.11穿透式電子顯微鏡(TEM)樣品製備......................22
3.12序列比對系統分析...................................22
3.13革蘭氏染色.........................................23
3.14細菌之溫度生理特性測試.............................23
3.15細菌之鹽度生理特性測試.............................23
3.16細菌之pH值耐受度生理特性測試.....................24
3.17 API 20 NE 生化反應................................24
肆、結果..................................................26
4.1 細菌的型態與特性...................................26
4.2 細菌的確認與親緣關係的比較.........................45
4.3細菌溫度之生理特性測試..............................49
4.4細菌鹽度生理特性測試................................51
4.5細菌酸鹼之生理特性測試..............................54
伍、討論..................................................57
5.1分離菌株與底質......................................57
5.2細菌生存環境........................................57
5.3菌株鑑定............................................58
陸、結論..................................................61
參考文獻.................................................62

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