印度梨型孢真菌促進小白菜生長相關基因之功能性探討

Abstract

印度梨型孢真菌 ( Piriformospora indica, P. indica ) 是一種新的根部內共生性真菌 ( root endophyte fungus )，屬於擔子菌門 ( Basidiomycoya )、蕈菌綱 ( Hymenomycetes )、梨型孢屬的真菌。已知對植物的生長發育有促進的作用，可以增加植物生物量 ( biomass )，也可以幫助宿主植物對抗逆境、增加抗氧化能力以及對病原菌具有抗性。本實驗以小白菜 ( Brassica campestris subsp. chinensis L. ) 作為與P. indica 共生的實驗植物，將其栽種於土壤中，P. indica可促進宿主小白菜根部的生長發育，藉使其地上部葉片與生物量皆大於未與P. indica共生之小白菜，除了促進小白菜生長，P. indica也可藉由還原態glutathione ( GSH ) 的提升來幫助小白菜對抗鹽、乾旱以及熱逆境。先前的研究已知P. indica對小白菜的生長促進作用是經由加強生長素 ( auxin ) 生合成與運輸、細胞膜轉運及根毛生長等各類型基因之表現而達成，本論文想進一步驗證這些基因在生長促進的功能，因此從先前所建立的扣減式cDNA基因庫中，選取促進生長相關之基因，如：AUX1、EXPA17、GSTU、PIN3、RHD4和V-ATPase，進行個別轉植於阿拉伯芥中，大量表現並分析阿拉伯芥之生理及型態變化。結果顯示，這些基因轉殖株分別顯現glutathione含量提升，促進植物的根部生長、根系密集或其地上部葉片生長旺盛，其中35S::BcGSTU 轉殖株亦可幫助植物提升對鹽和乾旱的耐受性。綜合結果，我們證明P. indica 促進小白菜誘導生長相關基因之表現後可促進小白菜生長，提升GSH的含量，並且藉由降低因為逆境而產生的H2O2累積來抵抗逆境，保護植物。

Piriformospora indica (P. indica) is a new root endophytic fungus belonging to the Hymenomycetes of the Basidiomycota. P. indica interacts with the roots of various mono- and dicotyledonous plants, showing the positive effect on biomass production. In addition to this growth promoting effect, P. indica also has the potential to induce resistance to fungal disease and to increase the antioxidative activities of plants. After the colonization of P. indica, plant becomes more tolerance to salt and drought. In this study, we used Chinese cabbage (Brassica campestris subsp. chinensis L.) for colonization with P. indica. After colonization for 7 days, six genes expression level of glutathione-S-transferase parC (BcGSTU), aux resistant 1 (BcAUX1) , efflux carrier, (BcPIN3), V-type proton ATPase (BcV-ATPase), expansin A17 (BcEXPA17),and root hair defective 4 (BcRHD4) were up-regulated and we have constructed above genes in pCAMBIA1300-35S-nos vector and tend to investigate gene function of the overexpressing cabbages. In glutathione assay, we found out the concentration of glutathione increased especially in leaves when Chinese cabbage colonized with P. indica. According to this result, we studied the tolerance Chinese cabbage colonized with P. indica under different stress. The results show that Chinese cabbage colonized with P. indica reduced the concentration of hydrogen peroxidase under NaCl stress indicated P. indica could enhance the salt tolerance of Chinese cabbage. We overexpressed these six gene in Arabidopsis and found out these gene could up-regulate the glutathione content and promote the growth of laterial roots, root length, and biomass. 35S::BcRHD4 could promote root hiars and 35S::BcGSTU could against the salt stress and drought sress. We figured out that glutathione play a role in plant growth and tolerance by P. indica colonized with Chinese cabbage.