臺灣博碩士論文加值系統

蘭菌菌種經分離與純化後，確認所分離之有益蘭菌屬於絲核菌屬，雙核R02菌株屬於AG-Bb，而多核R04菌株屬於AG-6。菌種繁殖建議可以使用2％ MP溶液。由本省各地所收集到不同金線連品系，在30/25℃下，品系代號C, P, L1, L3及T之存活率皆在70％以上。利用電解質滲漏法以50℃相對傷害值，可作為台灣金線連進行耐熱篩選之重要指標。另將植株置於埔里及台大溫室，以生長於埔里較台大溫室的鮮重較重。
台灣金線連種子共生發芽試驗，以燕麥培養基接種蘭菌（R02）於60天後，發芽率為80-86％。培養基覆蓋濾紙及接種蘭菌，可促進台灣金線連種子發芽及後續生長。瓶內或瓶外接種蘭菌後都可顯著促進植株的生長，並增加植株的葉綠素含量，使用塑膠袋栽培可達到省時、省工及減少農藥施用等目的。利用顯微鏡可觀察到菌絲進入根部後，菌絲互相纏繞，形成菌絲團塊等，屬於菌球消化之感染模式。台灣金線連接種蘭菌之菌根植株，根部同時具有酸性及鹼性磷酸酶，但未接種蘭菌之非菌根植株，則只有酸性磷酸酶。以根、莖及葉進行酵素活性與成分分析，植株同時具有過氧化酶、酸性磷酸酶及CuZnSOD與MnSOD二種超氧歧化酶，另外測定SOD、多醣、多酚類化合物、維生素C及磷酸根離子含量分析，接種蘭菌皆較未接種者，具有較高之含量。
彩葉蘭種子發芽以使用燕麥培養基並接種蘭菌R02，於播種後60天發芽率為65.6％較佳。以顯微鏡觀察種子發芽過程，可觀察到種子由吸水膨大、突破種皮、形成原球體一直到長出葉原體等不同階段的發育情形，及菌種侵入細胞之感染情形，利用組織化學測定，接種蘭菌之原球體，含有脂質、蛋白質及不可溶性多醣等成分，但對照組僅有少許之脂質及不可溶性多醣。植株無論於瓶內或瓶外接種蘭菌，皆可顯著促進植株生長，在彩葉蘭接種試驗，推薦蘭菌代號R01比R02的菌株為佳。若將已具花芽分化的植株置於人工氣候室，則以30/25℃的花梗抽長較快，利用顯微鏡可觀察到花芽發育與人工授粉後整個種子發育過程，果莢於授粉後35天開始變色，建議授粉後40-45天為種子之最佳採收期。
接種蘭菌R01、R02及R04可促進雜交種種子發芽及種苗與植株的生長，但F1雜交種的種子則無法發芽。30/25℃下雜交種植株的生長勢較25/20℃及20/15℃為佳，且接種蘭菌之菌根植株在四種溫度下，都較未接種之非菌根植株為佳，且接種蘭菌可增加植株的葉綠素含量。雜交種形態特徵介於台灣金線連與彩葉蘭之間，對於病蟲害的危害不具抗性。

Abstract
The isolated orchid mycorrhizal fungi (OMF) were cultured and purified. The OMF used for inoculation belonged to Rhizoctonia spp. Binucleate OMF R02 belongs to AG-Bb, while multinucleate R04 belongs to AG-6. Using 2% of MP solution could produce the highest dry weight of OMF,s ( R02 ) hypha. Those heat-resistant lines of Anoectochilus formosanus Hayata previously collected from various parts of Taiwan by our laboratory were cultivated in the phytoton at NTU for further study. However, survival rate of the five lines （C, P, L1, L3 and T）of A. formosanus at 30/25℃ reached 70%. Heat tolerance of A. formosanus was evaluated with injury values of cell membrance thermostability at 50℃ and this could be used as importment heat tolerance index. Five lines of A. formosanus grew in Puli had higher fresh weight than those at NTU.
Seeds inoculated with R02 OMF exhibited 80-86% of germination percentage 60 days in vitro on oat meal agar medium (OMA). Put filter paper on agar medium and the inoculation of OMF could promote seed germination and growth for this medical use of orchid. Effects of OMF in vitro or in vivo on the growth of A. formosanus were conducted. Results showed that A. formosanus inoculated with fungi could enhance growth and increase chorophyll contents. Plant cultured in plastic bag showed enhanced plant growth. Thus less pesticide or insecticide was needed. The fungal hypha penetrated cortex cells and formed pelotons was observed by microscopy, and results showed that was tolypophagy type of infection. Enzymatic histochchemistry test on roots of non-mycorrhizal control and inoculated with OMF were compared. The mycorrhizal root had both activities of acid and alkaline phosphatases, and non-mycorrhizal root had only acid phosphatase activity in A. formosanus. Enzyme activity and content anaysis of peroxidase, acid phosphatase, CuZnSOD and MnSOD activities were compared among root, stem and leaf and results showed that they were present in both non-mycorrhizal and OMF. SOD activity was higher in mycorrhizal than non-mycorrhizal control. Contents of polyphenol, polysaccharides, ascorbic acid and phoshate of A. formosanus were significantly higher in mycorrhizal plants.
In vitro germination percentage of Haemaria discolor var. dawsoniana was the highest（65.6%）for those seeds inoculated with R02 on OMA that exhibited germination percentage for 60 days. The developmental processes of germinating seeds were observed by microscopies. Seed germination occurred after water uptake, and seed coat was ruptured by embryo. Then papilla, protocorm, apical meristem appeared. Symbiotic germination showed that OMF hypha infected root cortex cells to form pelotons intracellularly. Enzymatic histochchemistry test showed that mycorrhizal protocorms contained mass lipid, protein and polysaccharide products, but non-mycorrhizal control only contained much less lipid and polysaccharide. The inoculation of OMF both in vitro or in vivo could enhance seedling development and growth of H. discolor. R01 was better than R02 for enhancing the growth of H. discolor. Before flower development, if plants were put in 30/25℃ day/night temperature in the phytoron, flower stalk length couble be increased. Microscopies were used to observe processes of flower and seed development. Cross section of capsule development 35 days after hand pollination seeds changed from white to brown color. It was suggested that hand pollination for 40-45 days, capsules of Haemaria discolor var. dawsoniana was the best harvest period.
Inoculate Rhizoctonia spp. (R01, R02 and R04 isolates) of orchid mycorrhizal fungi (OMF) could enhance seed germination, seedling and plantlet growth of Haemaria discolor var. dawsoniana×Anoectochilus formosanus Hayata, but resulted in no seed germination for F1 hybrid seeds. Plant at 30/25℃ showed that better growth than 25/20℃ and 20/15℃. Under the three studied temperature, the inoculation of OMF resulted in better growth than non-mycorrhizal control. Chorophyll content of Haemaria discolor var. dawsoniana × Anoectochilus formosanus Hayata increased as inoculated with OMF. Morphology of hybrid plant changed. The Haemaria discolor var. dawsoniana × Anoectochilus formosanus Hayata plant showed no disease and insects resistance.

目錄
中文摘要‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧Ⅰ
英文摘要‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧Ⅲ
第一章、前言
研究目‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧ 1
台灣金線連之繁殖、栽培管理、生理及藥理活性分析‧‧‧‧‧‧‧ 2
彩葉蘭的繁殖與栽培管理‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧ 10
蘭菌的分離、純化、鑑定與應用‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧14
第二章、蘭菌（絲核菌屬）的分離、純化、鑑定與繁殖
中文摘要‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧31
前言‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧31
材料與方法‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧32
結果與討論‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧35
結論‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧38
參考文獻‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧39
圖表‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧41
英文摘要‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧48
第三章、台灣金線連之耐熱性篩選、菌根生理、酵素活性及成分分析
中文摘要‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧49
前言‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧49
材料與方法‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧53
結果與討論‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧61
結論‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧74
參考文獻‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧76
圖表‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧83
英文摘要‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧ 102
第四章、彩葉蘭和彩葉蘭與台灣金線連雜交種之種子發芽、種苗生長、花期調節及組織化學檢定
中文摘要‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧ 103
前言‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧ 104
材料與方法‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧ 106
結果與討論‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧ 112
結論‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧ 126
參考文獻‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧ 127
圖表‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧ 131
英文摘要‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧ 163
第五章、總結‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧ 164

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