水稻清流對於瘤野螟與褐飛蝨相關防禦反應

Plant Defensive Response of Rice Variety Qingliu against Cnaphalocrocis medinalis and Nilaparvata lugens

李益(Yi Li)

指導教授 ： 莊汶博

國立臺灣大學/生物資源暨農學院/農藝學研究所/碩士(2019年)

https://doi.org/10.6342/NTU201903921

摘要

水稻為亞洲地區主要糧食作物。水稻在栽培過程中需面對諸多挑戰，其中蟲害為影響產量因素之一，未嚴加防範可影響近兩成產量。面對不同口器的害蟲，植物也對此演化出不同的植物防禦機制因應。然而因植物防禦機制複雜且耗能，故鮮少有植物能同時具備對不同口器的昆蟲有抗蟲能力。據前人研究水稻種原抗蟲結果中，我們得到了一個具備雙重抗性潛力的水稻品種「清流」，但對於其防禦模式尚未明瞭，因此本實驗以刺吸式口器的褐飛蝨及咀嚼式口器的瘤野螟作為蟲害處理，以感蟲品種台中在來一號(TN1)作為對照品種。首先透過生物檢定檢驗其抗蟲能力，發現清流均能顯著降低褐飛蝨與瘤野螟的攝食量及相對生長速率，MSST的結果也顯示在清流水稻半成株期也能保有對於褐飛蝨的抗性。使用RNA定序及測定清流防禦相關基因的表現量，比較不同口器害蟲危害及抗、感性品種間轉錄體學層次的差異性，發現清流在直接受到害蟲攻擊部位普遍有較多顯著調升的基因，同時也受不同口器的昆蟲攻擊而誘導不同防禦相關基因，瘤野螟會誘導protease、proteinase inhibitor等影響昆蟲腸道消化的調控基因或是與植物荷爾蒙JA生合成相關的lipoxygenase (LOX)、jasmonate-induced protein(JIP)基因，而褐飛蝨則誘導與癒傷葡聚醣(callose)沉積相關的β-1,3-glucanase、glucanase等基因，顯示不同口器的昆蟲誘導的植物防禦基因也不同。同時，在測定SA及JA-Ile這兩個相關荷爾蒙與苯丙胺酸裂解酶(PAL)的酵素活性中，發現清流對於防禦機制相關酵素(PAL)活性的提升速度較感蟲品種更快速或是更持久，PAL相關基因的表現量趨勢更是符合PAL酵素的變化趨勢。藉本實驗結果初步構築清流此品種雙重抗性水稻的防禦模式，相較感蟲品種TN1而言，其更能夠有效且快速地轉錄下游產物來啟動防禦，同時能維持較長時間的防禦期，而從RNA定序結果中，也有諸多針對不同口器害蟲的防禦相關基因被大量誘導表現，未來可供進一步的檢驗使用。

關鍵字

抗蟲檢定 ； RNA定序 ； 植物荷爾蒙 ； PAL ； 刺吸式口器 ； 咀嚼式口器

並列摘要

Rice (Oryza sativa L.) is the most important staple food in Asia. During rice cultivation, plants would face to several environmental and biotic challenges, such as insect infestation. Insect infestation would affect around 20% yield loss in the world. Plants have evolved different defense mechanisms to detect insect infestation based on the insect mouthparts. However, the plant defense mechanism is very sophisticated and costs lots of energy. Thus, very few plants would carry multi-insect resistant traits against insects with different monthparts. Based on our previous study, we found one rice variety, Qingliu, which would be the candidate for the dual-insect resistant variety against two different insect feeding guilds. However, it is unclear how Qingliu defend two feeding insect feeding guilds. Thus, the goal of this study is understanding the defense mechanism of Qingliu. First, through bioassays, we found Nilaparvata lugens would produce few honeydew and Cnaphalocrocis medinalis larvae would have lower relative growth rate on Qingliu. Based on MSST data, it showed that Qingliu would have resistant ability against N. lugens during mid-vegetative stage. In addition, we measured the transcriptome with two insect infestation on Qingliu and susceptible check variety, TN1, by RNA-sequencing and further validated the gene expression pattern. The results indicated that more defensive-related genes in Qingliu were induced under herbivore infestation. However, there is different expression pattern in Qingliu under different insect feeding guilds. Under C. medinalis infestation, protease & proteinase inhibitor related genes, lipoxygenase, JA-biosynthetic gene, and jasmonate-induced protein were induced in Qingliu. However, under N. lugens infestation, β-1,3-glucanase, callose-related genes, and glucanase were induced in Qingliu. In addition, two phytohormone (SA and JA-Ile) and phenylalanine ammonia lyase (PAL) enzyme activity were measure. We found the enzyme activity of PAL in Qingliu was faster induced or maintained its activity longer. Furthermore, PAL-related gene expression pattern was positive correlated with the enzyme activity pattern. Based on this study, we could establish the defense pattern of Qingliu against two insect pests. Comparing with susceptible rice variety, TN1, Qingliu would trigger downstream defensive genes faster and have a longer defensive period. From the result of RNA-seq, we found lots of plant defensive-relative genes with insect monthpart specific were highly induced. These results would be validated in the future.

並列關鍵字

Insect-resistant Bioassay ； RNA Sequencing ； Phytohormone ； PAL ； Piercing-sucking Mouthparts ； Chewing Mouthparts

參考文獻

Alvarez A, Montesano M, Schmelz E & Ponce de León I (2016) Activation of shikimate, phenylpropanoid, oxylipins, and auxin pathways in Pectobacterium carotovorum elicitors-treated moss. Frontiers in Plant Science 7: 328.

Auclair JL (1958) Honeydew excretion in the pea aphid, Acyrthosiphon pisum (Harr.)(Homoptera: Aphididae). Journal of Insect Physiology 2: 330-337.

Awmack CS & Leather SR (2002) Host plant quality and fecundity in herbivorous insects. Annual Review of Entomology 47: 817-844.

Bell E & Mullet JE (1993) Characterization of an Arabidopsis lipoxygenase gene responsive to methyl jasmonate and wounding. Plant Physiology 103: 1133-1137.

Bodenhausen N & Reymond P (2007) Signaling pathways controlling induced resistance to insect herbivores in Arabidopsis. Molecular Plant-Microbe Interactions 20: 1406-1420.