植物常會遭受不同的生物及非生物性逆境的危害，然而當植物遇到環境逆境時，無法立即移動脫逃，需要仰賴體內啟動一連串的防禦機制，來達到自我保護的功能。而屬生物性逆境中的植物病蟲害，一直是造成全球農作物產量及品質受損之重要因素；近年來，利用植物遺傳工程以培育抗逆境新作物品種，已成為常被應用且有效的病害管理方法。近幾年科學家利用篩選突變株發現NPR1 (nonexpresser of PR genes)，為一在植物啟動系統性誘導抗病機制時重要的調控蛋白質。本實驗室先前培育出大量表現阿拉伯芥NPR1 (AtNPR1 ) 之番茄轉殖品系 (CL5915)，並將其對八種重要之番茄病害的抗病性作測試，結果證實，這些轉殖品系具有廣效抗病性，特別對維管束病害及葉部病害，然而其抗病機制至今仍不清楚。本論文針對特定AtNPR1轉殖品系，以青枯病菌為研究對象，分析其分子抗病機制，並評估其園藝性狀。實驗結果發現，在轉殖品系中，葉部組織之特定PR基因表現量確實較未轉殖品系高，推測此為轉殖品系對葉部病害有產生抗性的原因之一；但根部組織之特定PR基因表現量和未轉殖品系無顯著差異，無法解釋AtNPR1轉殖品系對於維管束病害之抗性。所以進一步利用cDNA微陣列技術分析大量基因，以探討AtNPR1轉殖品系可能誘導之抗病機制。進一步分析及比對微陣列結果發現，AtNPR1轉殖品系有部分基因會共同受青枯病菌與過量銅誘導表現，推測這兩個防禦訊息間有相互交集。而本論文也進一步將番茄植物處理園藝上常見的非生物性逆境，證實未轉殖及轉殖品系對非生物性逆境的反應耐受性一致，並沒有因為基因轉殖而造成性狀上的缺失。由於現有的轉殖品系 (CL5915) 對於番茄嚴重的病毒病害的抗性沒有顯著提高，所以本論文也將AtNPR1轉殖入一個對番茄黃葉捲曲病毒有中度抗性的番茄品種 (CLN2116B)，並已成功培育具抗病性的新番茄品系，預期將來可進一步應用於栽培品種，當作抗病育種之抗病性狀來源，以傳統育種雜交法增加其抗病範圍，有效達成作物病害管理防治目的。 Abstract In nature, crop plants constantly encounter various biotic and abiotic stresses, which can severely affect agricultural productivity. Pathogens, particularly, are limiting factors reducing crops quality and quantity seriously. Genetic engineering of disease-resistance through transferal of plant defense-related genes into crop is a valuable disease-control approach. Among the defense genes used to genetically engineer systemic acquired resistance in plants, Arabidopsis NPR1 (nonexpresser of PR genes) is of particular interest for its being a central regulator of plant defense responses. The exploration of empolying Arabidopsis NPR1 (AtNPR1) gene for genetics engineering disease-resistance in tomato plants have led to the production of transgenic tomato lines conferring broad-spectrum disease-resistance, especially to vascular and leaf pathogens. In this work, the possible resistance mechanism employed by a selected AtNPR1 transgenic line and its interactions with Ralstonia solanacerarum were further studied. The results showed that some PR (pathogenesis-related) genes were constitutively expressed at a high level in the leaf tissues of the transgenic plants and thus may account for the enhanced resistances to the leaf pathogens. By further employing cDNA microarray approach, using customized tomato cDNA microarray, a group of putative AtNPR1 overexpression-induced genes were identified and a model for defense mechanism is proposed. By cross-referencing these data with the microarray data obtained from other abiotic stress studies, possible cross-links between biotic stress and heavy metal stress response were revealed. Evaluation of the horticulture traits of the transgenic tomato showed that the transgenic plants responded similarity, as did the wild type plants under drought and salinity stress. Furthermore, because none of the transgenic lines generated in the genetic background of CL5915 tomato cultivar conferred enhanced resistance to virus diseases, a tomato cultivars, CLN2116B, which carries the resistance trait to tomato yellow leaf curl virus was used as the background cultivar for transformation with AtNPR1. New CLN2116B transgenic plants were selected and characterized. Molecular analyzes revealed that expression of some PR genes was constitutively enhanced in these transgenic plants, suggesting a potential enhanced of disease-resistance in these transgenic plants.
