深海和南極產(chǎn)油真菌篩
發(fā)布時(shí)間：2018-07-25 10:04
【摘要】：本論文的內(nèi)容主要是研究大洋與南極樣品的真菌多樣性、采用尼羅紅與氯化三苯基四氮唑(TTC)方法進(jìn)行產(chǎn)油真菌的初篩、采用氣相色譜方法對產(chǎn)油菌的脂肪酸成分進(jìn)行分析及研究培養(yǎng)基成份對產(chǎn)油菌產(chǎn)油發(fā)酵的影響。主要結(jié)果如下:對中太平洋(6個(gè)站位)、南大西洋(2個(gè)站位)及西太平洋(22個(gè)站位)共30個(gè)站位的沉積物進(jìn)行分離鑒定,一個(gè)獲得75株真菌,通過ITS鑒定結(jié)果顯示,這75株菌屬于14個(gè)屬,它們分別是:Aspergillus sp.(27株)、Penicillium sp.(21株)、Cladosporium sp.(8株)、Fusarium sp.(6株)、Alternaria(2株)、Exophiala sp.(2株)、Engyodontium sp.(2株)、Acremonium sp.(1株)、Nectria sp.(1株)、Tritirachium sp.(1株)、Aureobasidium sp.(1株)、Thielavia sp.(1株)、Lecythophora sp.(1株)、Sarocladium sp.(1株)。三個(gè)海區(qū)中菌株數(shù)量最多的為曲霉屬的真菌,可見曲霉屬真菌在這三個(gè)海區(qū)中為優(yōu)勢屬,其次為青霉屬。在太平洋中,枝孢屬的真菌也有一定的分布。其余屬均只有幾株,表明在三個(gè)大洋中分布較少。同時(shí)分別使用尼羅紅和TTC對大洋樣品中分離得到的369株真菌進(jìn)行產(chǎn)油初篩,一共獲得61株產(chǎn)油菌,分別屬于10個(gè)屬:Aspergillus(21株)、Fusarium(13株)、Cladosporium(10株)、Penicillium(9株)、Engyodontium(2株)、Sarocladium(2株)、Alternaria(1株)、Aureobasidium(1株)、Lecythophora(1株)、Nectria(1株)。產(chǎn)油菌所占比例為18.70%。其中曲霉屬菌株占產(chǎn)油菌比例最高,為30.43%,其次為鐮刀屬菌株、枝孢屬菌株與青霉素菌株,分別占總產(chǎn)油菌屬的18.84%、14.49%與13.04%。對中國第29次南極科學(xué)考察獲得的南極普里茲灣海域23個(gè)站位的深海沉積物樣品進(jìn)行了真菌分離培養(yǎng),共獲得了11個(gè)屬的21株真菌。分別是:Penicillium(9株)、Cladosporium(3株)、Acremonium(1株)、Aureobasidium(1株)、Botryotinia(1株)、Eurotiales(1株)、Exophiala(1株)、Fusarium(1株)、Nectria(1株)、Phoma(1株)、Sarocladium(1株)。南極普里茲灣海域的真菌多樣性較高,其中青霉屬菌株最多,占總菌株比例為42.85%,是普里茲灣真菌中的優(yōu)勢屬,其次為枝孢屬菌株,占總菌株比例為14.29%。分別用尼羅紅和TTC對南極普里茲灣海域分離得到的21株真菌進(jìn)行產(chǎn)油初篩,共獲得8株產(chǎn)油菌,分別屬于7個(gè)屬:Penicillium(2株)、Aureobasidium(1株)、Cladosporium(1株)、Eurotiales(1株)、Exophiala(1株)、Fusarium(1株)、Sarocladium(1株)。產(chǎn)油菌占總菌株的比例為38%。對分離得到的產(chǎn)油菌進(jìn)行脂肪酸分析,結(jié)果表明:這些產(chǎn)油菌所含的基本為C16-C18脂肪酸,不飽和脂肪酸含量較高,脂肪酸中具有較長的碳直鏈且沒有碳支鏈,因此可以作為生物柴油制備的原料油�？紤]到油脂生產(chǎn)成本,尋找可利用廉價(jià)底物高產(chǎn)油脂的微生物具有深遠(yuǎn)的意義。為了尋找能夠直接利用廉價(jià)底料作為碳源產(chǎn)油的真菌菌株,本文使用含有TTC的糖蜜培養(yǎng)基分離南極沉積物樣品,獲得一株產(chǎn)油菌,為P7-07-M1,屬名為:Nectria,該菌株可以直接利用糖蜜作為碳源發(fā)酵產(chǎn)油。對從南極產(chǎn)油菌中選擇一株產(chǎn)油菌(P7-07-M1),從大洋產(chǎn)油菌中選擇兩株產(chǎn)油菌(JL7KGC01A-M7與MP4CTD-11-M5)進(jìn)行培養(yǎng)基優(yōu)化,結(jié)果顯示:菌株P(guān)7-07-M1達(dá)到最高產(chǎn)油量的時(shí)間為第8天,最適氮源和最適碳源分別為蛋白胨與葡萄糖,最適C/N比為168,最佳Cu SO4.5H2O濃度為0.0001g/L,最佳Mn SO4.H2O濃度為0.004g/L,通過培養(yǎng)基優(yōu)化,菌株P(guān)7-07-M1產(chǎn)油量提升了27.89%;菌株JL7KGC01A-M7達(dá)到最高產(chǎn)油量的時(shí)間為第7天,最適氮源和最適碳源分別為蛋白胨與葡萄糖,最適C/N比為140,最佳Cu SO4.5H2O濃度為0.0002g/L,最佳Mn SO4.H2O濃度為0.003g/L,通過培養(yǎng)基優(yōu)化,菌株JL7KGC01A-M7的產(chǎn)油量提升了30.15%;菌株MP4CTD-11-M5達(dá)到最高產(chǎn)油量的時(shí)間為第7天,最適氮源和最適碳源分別為蛋白胨與葡萄糖,最適C/N比為140,最佳Cu SO4.5H2O濃度為0.0001g/L,最佳Mn SO4.H2O濃度為0.004g/L,通過培養(yǎng)基優(yōu)化,菌株MP4CTD-11-M5的產(chǎn)油量提升了111.16%。菌株MP4CTD-11-M5培養(yǎng)基優(yōu)化效果最顯著。
[Abstract]:The main content of this paper is to study the fungal diversity of Oceanic and Antarctic samples. Using Nile red and three phenyl tetrazolium chloride (TTC) method to screen the oil producing fungi, the fatty acid composition of the oil producing bacteria was analyzed by gas chromatography and the influence of the medium on the production of oil producing bacteria was studied. The main results are as follows: The sediments of 30 stations in the central Pacific (6 stations), southern the Atlantic (2 stations) and the Western Pacific (22 stations) were isolated and identified, and 75 fungi were obtained. The results of ITS identification showed that the 75 strains belonged to 14 genera: Aspergillus sp. (27 strains), Penicillium sp. (21 strains), Cladosporium sp. (8 strains), Fusarium sp. (6 strains), Alternaria (2 strains), Exophiala sp. (2 strains), Engyodontium sp. (2 strains), Acremonium sp. (1 strains), Nectria sp. (1 strains), Tritirachium sp. (1 strains), Aureobasidium 1, 1 plants, 1 strains (1 strains). The largest number of strains of the genus Aspergillus in the three sea areas, visible Aspergillus fungi In the three sea areas, the second is Penicillium. In the Pacific, the fungi of the genus cladospora are also distributed. The other genera are only a few, indicating that there are only a few strains in the three oceans. At the same time, 369 strains of fungi isolated from the ocean samples were screened using Nile red and TTC respectively. A total of 61 strains of oil producing bacteria were obtained. Don't belong to 10 genera: Aspergillus (21 strains), Cladosporium (13 strain), Penicillium (10 strain), Penicillium (9 strain), Engyodontium (2 strain), Sarocladium (2 strain), Alternaria (1 strain), Aureobasidium (1 strain), Lecythophora (1 strain), Nectria (1 strain). The proportion of oil producing bacteria is 18.70%., and the proportion of Aspergillus is the highest, 30.43%, and second is sickle. Spore strains, cladospora strains and penicillin were 18.84%, 14.49% and 13.04%., respectively, and 14.49% of the deep-sea sediment samples from 23 stations in the Antarctic Bay of Antarctica were isolated and cultured by 13.04%., and 21 fungi of 11 genera were obtained, respectively: Penicillium (9 strains), Cladospo Rium (3 strains), Acremonium (1 strain), Aureobasidium (1 strain), Botryotinia (1 strain), Eurotiales (1 strain), Exophiala (1 strain), Fusarium (1 strain), Nectria (1 strain), Phoma (1 strain), Sarocladium (1). The fungal diversity in the sea area of Antarctica is higher, among which Penicillium is most, and the proportion of total strains is 42.85%. It is the dominant genera of the Prem Bay fungi. The second was cladospora, and the proportion of total strains was 14.29%., 21 strains of fungi isolated from the sea area of Antarctica were screened with Nile red and TTC respectively, and 8 strains of oil producing bacteria were obtained, which belong to 7 genera: Penicillium (2 strains), Aureobasidium (1 strains), Cladosporium (1 strains), Exophiala (1 strains), Fusarium (1 strains), S, S, S. Arocladium (1 strains). The ratio of the oil producing bacteria to the total strains was 38%. to analyze the fatty acid of the isolated oil producing bacteria. The results showed that the oil producing bacteria contained basically C16-C18 fatty acids, high content of unsaturated fatty acids, long carbon chain and no carbon branching in fatty acids, so they could be used as raw materials for biodiesel. Oil. Considering the cost of oil production, it is of great significance to find the microorganisms which can make use of cheap substrates to produce high yield oil. In order to find a fungal strain which can be used as a carbon source directly using cheap substrate, this paper uses a molasses medium containing TTC to separate Antarctic sediment samples and obtain a strain of P7-07-M1, named Nectria. The strain can be fermented with molasses as a carbon source to produce oil. A strain of oil producing bacteria (P7-07-M1) from the Antarctic oil producing bacteria (P7-07-M1) is selected, and two strains of oil producing bacteria (JL7KGC01A-M7 and MP4CTD-11-M5) are selected from the marine oil producing bacteria to optimize the medium. The results show that the strain P7-07-M1 reaches the maximum oil yield for eighth days, the optimum nitrogen source and the optimum. The optimum C/N ratio was 168, the optimum Cu SO4.5H2O concentration was 0.0001g/L and the optimum Mn SO4.H2O concentration was 0.004g/L. The oil production of strain P7-07-M1 increased by 27.89% by the optimization medium, and seventh days for the strain JL7KGC01A-M7 to reach the highest oil production, and the optimum nitrogen source and the optimum carbon source were peptone and Portuguese. The optimum C/N ratio was 140, the optimum Cu SO4.5H2O concentration was 0.0002g/L, the optimum Mn SO4.H2O concentration was 0.003g/L, the oil production of strain JL7KGC01A-M7 increased by 30.15%, the strain MP4CTD-11-M5 reached the maximum oil production time seventh days, the optimum nitrogen source and the optimum carbon source were peptone and glucose, and the optimum C/N ratio was 140. The optimum Cu SO4.5H2O concentration is 0.0001g/L and the optimum Mn SO4.H2O concentration is 0.004g/L. The optimization of the oil production of the strain MP4CTD-11-M5 improves the optimization effect of the MP4CTD-11-M5 medium of 111.16%. strain by optimizing the medium of the strain MP4CTD-11-M5.
【學(xué)位授予單位】：國家海洋局第三海洋研究所
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2015
【分類號】：TQ929;Q93;TE667
本文編號：2143462
[Abstract]:The main content of this paper is to study the fungal diversity of Oceanic and Antarctic samples. Using Nile red and three phenyl tetrazolium chloride (TTC) method to screen the oil producing fungi, the fatty acid composition of the oil producing bacteria was analyzed by gas chromatography and the influence of the medium on the production of oil producing bacteria was studied. The main results are as follows: The sediments of 30 stations in the central Pacific (6 stations), southern the Atlantic (2 stations) and the Western Pacific (22 stations) were isolated and identified, and 75 fungi were obtained. The results of ITS identification showed that the 75 strains belonged to 14 genera: Aspergillus sp. (27 strains), Penicillium sp. (21 strains), Cladosporium sp. (8 strains), Fusarium sp. (6 strains), Alternaria (2 strains), Exophiala sp. (2 strains), Engyodontium sp. (2 strains), Acremonium sp. (1 strains), Nectria sp. (1 strains), Tritirachium sp. (1 strains), Aureobasidium 1, 1 plants, 1 strains (1 strains). The largest number of strains of the genus Aspergillus in the three sea areas, visible Aspergillus fungi In the three sea areas, the second is Penicillium. In the Pacific, the fungi of the genus cladospora are also distributed. The other genera are only a few, indicating that there are only a few strains in the three oceans. At the same time, 369 strains of fungi isolated from the ocean samples were screened using Nile red and TTC respectively. A total of 61 strains of oil producing bacteria were obtained. Don't belong to 10 genera: Aspergillus (21 strains), Cladosporium (13 strain), Penicillium (10 strain), Penicillium (9 strain), Engyodontium (2 strain), Sarocladium (2 strain), Alternaria (1 strain), Aureobasidium (1 strain), Lecythophora (1 strain), Nectria (1 strain). The proportion of oil producing bacteria is 18.70%., and the proportion of Aspergillus is the highest, 30.43%, and second is sickle. Spore strains, cladospora strains and penicillin were 18.84%, 14.49% and 13.04%., respectively, and 14.49% of the deep-sea sediment samples from 23 stations in the Antarctic Bay of Antarctica were isolated and cultured by 13.04%., and 21 fungi of 11 genera were obtained, respectively: Penicillium (9 strains), Cladospo Rium (3 strains), Acremonium (1 strain), Aureobasidium (1 strain), Botryotinia (1 strain), Eurotiales (1 strain), Exophiala (1 strain), Fusarium (1 strain), Nectria (1 strain), Phoma (1 strain), Sarocladium (1). The fungal diversity in the sea area of Antarctica is higher, among which Penicillium is most, and the proportion of total strains is 42.85%. It is the dominant genera of the Prem Bay fungi. The second was cladospora, and the proportion of total strains was 14.29%., 21 strains of fungi isolated from the sea area of Antarctica were screened with Nile red and TTC respectively, and 8 strains of oil producing bacteria were obtained, which belong to 7 genera: Penicillium (2 strains), Aureobasidium (1 strains), Cladosporium (1 strains), Exophiala (1 strains), Fusarium (1 strains), S, S, S. Arocladium (1 strains). The ratio of the oil producing bacteria to the total strains was 38%. to analyze the fatty acid of the isolated oil producing bacteria. The results showed that the oil producing bacteria contained basically C16-C18 fatty acids, high content of unsaturated fatty acids, long carbon chain and no carbon branching in fatty acids, so they could be used as raw materials for biodiesel. Oil. Considering the cost of oil production, it is of great significance to find the microorganisms which can make use of cheap substrates to produce high yield oil. In order to find a fungal strain which can be used as a carbon source directly using cheap substrate, this paper uses a molasses medium containing TTC to separate Antarctic sediment samples and obtain a strain of P7-07-M1, named Nectria. The strain can be fermented with molasses as a carbon source to produce oil. A strain of oil producing bacteria (P7-07-M1) from the Antarctic oil producing bacteria (P7-07-M1) is selected, and two strains of oil producing bacteria (JL7KGC01A-M7 and MP4CTD-11-M5) are selected from the marine oil producing bacteria to optimize the medium. The results show that the strain P7-07-M1 reaches the maximum oil yield for eighth days, the optimum nitrogen source and the optimum. The optimum C/N ratio was 168, the optimum Cu SO4.5H2O concentration was 0.0001g/L and the optimum Mn SO4.H2O concentration was 0.004g/L. The oil production of strain P7-07-M1 increased by 27.89% by the optimization medium, and seventh days for the strain JL7KGC01A-M7 to reach the highest oil production, and the optimum nitrogen source and the optimum carbon source were peptone and Portuguese. The optimum C/N ratio was 140, the optimum Cu SO4.5H2O concentration was 0.0002g/L, the optimum Mn SO4.H2O concentration was 0.003g/L, the oil production of strain JL7KGC01A-M7 increased by 30.15%, the strain MP4CTD-11-M5 reached the maximum oil production time seventh days, the optimum nitrogen source and the optimum carbon source were peptone and glucose, and the optimum C/N ratio was 140. The optimum Cu SO4.5H2O concentration is 0.0001g/L and the optimum Mn SO4.H2O concentration is 0.004g/L. The optimization of the oil production of the strain MP4CTD-11-M5 improves the optimization effect of the MP4CTD-11-M5 medium of 111.16%. strain by optimizing the medium of the strain MP4CTD-11-M5.
【學(xué)位授予單位】：國家海洋局第三海洋研究所
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2015
【分類號】：TQ929;Q93;TE667
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