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研究生:江惟
研究生(外文):Jiang,Wei
論文名稱:分析臺灣產固氮酵母菌之增進植物生長能力之效益
論文名稱(外文):Characterization of nitrogen fixation yeast strains in Taiwan for growth promotion of plants
指導教授:傅士峰
指導教授(外文):Fu,Shih-Feng
口試委員:傅士峰周睿鈺陳威臣
口試日期:2021-08-06
學位類別:碩士
校院名稱:國立彰化師範大學
系所名稱:生物學系
學門:生命科學學門
學類:生物學類
論文種類:學術論文
論文出版年:2021
畢業學年度:109
語文別:中文
論文頁數:56
中文關鍵詞:
外文關鍵詞:
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植物在土壤中吸收的營養素以氮、磷、鉀最多。其中氮對植物的生長發
育占了非常重要的地位,氮不僅是細胞中的蛋白質、核酸和磷脂質的構成
元素之一,對植物體的光合作用、能量供應等方面也是不可缺少的,植物
從土壤吸收利用以 NH4
+、NO3為主,而土壤中氮元素來源來自於空氣中的
氮氣,少數由閃電固定,但絕大部分的氮元素會經由土壤中的固氮菌透過
固氮酶將 N2轉換成植物可利用的形式。在種植植物時會添加肥料植物生長
更為良好,而目前常用的肥料種類是化學肥料,大量使用化肥去滿足植物
的需求,將會導致土壤汙染、破壞土壤的蓄水能力、肥力,且土壤養分比例
被大幅度改變。有文獻指出,通常施用的化肥中,植物只能吸收 10%~40%。
農業需要與環境友善的可持續性種植,因此化肥的安全替代品為生物肥料。
目前常見的氮生物肥料大多屬於原核生物肥料,對於真核生物肥料研究較
少,但已有研究證實部分酵母菌同樣擁有固氮能力,而對台灣本土的酵母
菌所知有限,本研究已在彰化及南投地區的番茄根圈選出具有固氮能力的
酵母菌,因此本研究將進一步探討酵母菌對於促進單子葉或雙子葉作物(如:
阿拉伯芥 Arabidopsis thaliana、水稻 Oryza sativa、小白菜 Brassica rapa
chinensis)生長效益。試驗結果顯示:所選用之酵母菌具固氮能力,且固氮
能力有所差異,其中編號 JYC 548 的銨含量經檢測後數值最高。另外在培
養基中可觀察到所測試之植物,以編號 JYC 468 酵母菌對於促進雙子葉植
物生長之效果最佳,在單子葉植物影響不顯著,而編號 JYC 536 酵母菌對
於植物無法促進生長,後續栽培試驗,則是 JYC461 對於植物成長促進效果
較優,並且於小白菜與阿拉伯芥中可觀察到相同之生長促進現象,此可證
實部分固氮酵母菌具有生物肥料之潛能。
The nutrients that plants absorb in the soil are the most nitrogen, phosphorus,
and potassium. Among them, nitrogen plays a very important role in the growth
and development of plants. Nitrogen is not only one of the constituent elements
of proteins, nucleic acids and phospholipids in cells, but also indispensable to the
photosynthesis and energy supply of plants. Soil absorption and utilization are
mainly NH4+ and NO3. The source of nitrogen in the soil comes from nitrogen
in the air. A few are fixed by lightning, but most of the nitrogen will be converted
into plants by nitrogen-fixing bacteria in the soil through nitrogenase. Available
forms. When planting plants, fertilizers are added to plants to grow better. At
present, the commonly used fertilizer types are chemical fertilizers. Large
amounts of chemical fertilizers are used to meet the needs of plants, which will
cause soil pollution, destroy the soil’s water storage capacity, fertility, and soil
nutrients. The ratio has been changed drastically. It is pointed out in the literature
that plants can only absorb 10% to 40% of chemical fertilizers commonly used.
Agriculture requires sustainable planting that is environmentally friendly, so a
safe alternative to chemical fertilizers is bio-fertilizer. At present, most of the
common nitrogen biological fertilizers are prokaryotic fertilizers. There are few
studies on eukaryotic fertilizers. However, some studies have confirmed that
some yeasts also have the ability to fix nitrogen, and the knowledge of yeasts in
Taiwan is limited. This research has been conducted in Changhua. Yeasts with
nitrogen-fixing ability were selected from the tomato rhizosphere in Nantou area.
Therefore, this study will further explore the benefits of yeasts on promoting the
growth of monocotyledonous or dicotyledonous crops (such as Arabidopsis
thaliana, Oryza sativa, Brassica rapa chinensis). . The test results show that the
selected yeasts have nitrogen-fixing ability, and the nitrogen-fixing ability is
different. Among them, the ammonium content of JYC 548 has the highest value
after testing. In addition, the tested plants can be observed in the culture medium.
Yeast No. JYC 468 has the best effect on promoting the growth of dicotyledonous
plants, but there is no difference in monocotyledonous plants. Yeast No. JYC 536
has the least effect on the growth of plants. Obviously, the follow-up cultivation
test showed that JYC461 has a better effect on plant growth promotion, and the
same growth promotion phenomenon can be observed in Chinese cabbage and
Arab mustard, which can prove that some nitrogen-fixing yeasts have the
potential of biological fertilizer.
目錄
中文摘要........................................................................................................................3
Abstract.........................................................................................................................4
致謝................................................................................................................................5
縮寫表..........................................................................................................................10
1. 前言 ........................................................................................................................ 11
1-1.氮元素對於植物之重要性 ...........................................................................................11
1-2.土壤中的氮型態...........................................................................................................12
1-3.植物氮的自主獲取途徑 ...............................................................................................13
1-4. 土壤微生物活動對於植物之影響 .............................................................................14
1-5. 生物肥料的選擇及生產考量 .....................................................................................15
1-6. 研究目的.....................................................................................................................17
2. 研究方法 ................................................................................................................18
2-1 植株選擇(Plant selection).............................................................................................18
2-2 酵母菌(yeast strains).....................................................................................................18
2-3 酵母菌培養(cultivation of yeasts) ................................................................................19
2-4 酵母菌破菌(Yeast Broken) ..........................................................................................22
2-4-1 液態培養後破菌............................................................................................22
2-4-2 固態培養後破菌............................................................................................22
2-5 培養基內種子接種(Planting in the medium)...............................................................23
2-5-1 水稻種子接種................................................................................................23
2-5-2 小白菜種子接種............................................................................................23
2-5-3 阿拉伯芥(AtPLDZ2)種子接種 .....................................................................24
2-6 土壤中植物生長測試(Plant growth in soil test)..........................................................24
2-6-1 水稻土壤測試................................................................................................24
2-6-2 小白菜土壤測試............................................................................................25
2-6-3 阿拉伯芥土壤測試........................................................................................25
2-7 酵母菌固氮能力檢測(Yeast nitrogen fixation ability test)..........................................25
2-7-1 酵母菌在 N-free 培養基中生長情況 ..........................................................25
2-7-2 NH4
+含量檢測 ...............................................................................................26
2-8 植物鮮重檢測(measurement of plant biomass)............................................................27
2-8-1 培養基中植物鮮重檢測................................................................................27
8
2-8-2 土壤種植植物鮮重檢測................................................................................27
2-9 酵母菌保存(preparation of the yeast stock)..................................................................28
3. 試驗結果 ................................................................................................................29
3-1 檢測酵母菌在無氮培養基中生長情形 .......................................................................29
3-2 檢測定時生長之酵母菌破菌後銨含量 .......................................................................29
3-3 檢測定量酵母菌破菌後銨含量 ...................................................................................30
3-4 定時固氮酵母菌影響阿拉伯芥在低氮培養基生長之鮮重結果分析........................31
3-5 定時固氮酵母菌影響小白菜在低氮培養基生長之鮮重結果分析............................32
3-6 定時固氮酵母菌影響水稻在低氮培養基生長之鮮重結果分析................................33
3-7 固氮酵母菌影響阿拉伯芥在培養土生長之鮮重結果分析 .......................................33
3-8 固氮酵母菌影響小白菜在培養土生長之鮮重結果分析 ...........................................34
3-9 固氮酵母菌影響水稻在培養土生長之鮮重結果分析 ...............................................35
3-10 定量固氮酵母菌影響植物在低氮培養基生長之鮮重結果分析..............................35
4. 討論 ........................................................................................................................37
4-1 比較不同植物在酵母菌提供的影響下之生長差異 ...................................................37
4-2 酵母菌內銨含量檢測與實際用於植物之效果差異 ...................................................38
4-3 土壤與培養基之植物受酵母菌影響之生長差異 ......................................................39
5. 結論 ........................................................................................................................40
6. 參考文獻 ................................................................................................................41
7. 論文結果圖 ............................................................................................................44
Fig. 1 N-free 培養基中酵母菌生長之表型結果................................................................44
Fig. 2 定時兩天酵母菌破菌後銨含量之結果 ..................................................................45
Fig. 3 定量酵母菌破菌後之銨含量結果 ..........................................................................46
Fig. 4 定時固氮酵母菌影響阿拉伯芥在低氮培養基之結果 ..........................................47
Fig. 5 定時固氮酵母菌影響小白菜在低氮培養基之結果 ..............................................48
Fig. 6 定時固氮酵母菌影響水稻在低氮培養基之結果 ...................................................49
Fig. 7 固氮酵母菌影響阿拉伯芥在培養土之結果 ...........................................................50
Fig. 8 固氮酵母菌影響小白菜在培養土之結果 ...............................................................51
Fig. 9 固氮酵母菌影響水稻在培養土之結果 ...................................................................52
9
Fig. 10 定量固氮酵母菌影響阿拉伯芥在低氮培養基之結果 .........................................53
Fig. 11 定量固氮酵母菌影響小白菜在低氮培養基之結果..............................................54
8. 附錄 ........................................................................................................................55
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