2012-03-012024-05-13https://scholars.lib.ntu.edu.tw/handle/123456789/650747摘要：食用菇類具有提昇免疫力、防止老化功能或抗癌的效果，是重要的天然藥食同源的食材。將傳統菇菌業產品與醫療保健相結合，可大幅增加菇類產品的附加價值，達到產業升級的目標。隨著現代生物技術的發展，菇菌不僅可以提供特定基因的來源，也可作為特定蛋白質生產的生物反應器。藉由功能性評估，如細胞及動物實驗的方法來驗證其醫藥功效；也可藉由育種和雜交的生物技術，進一步自篩選出的下一代菌種中培育出具特定功能與療效的新品種並註冊其智慧財產權，而後配合其栽種技術的建立將之移轉於農民生產栽種。以菇菌作為異源蛋白質表達系統，更是現代生物技術在分子農場的新的突破。然而，目前轉基因菇的篩選多仰賴抗生素抗性基因，如潮黴素等。抗生素抗性基因的使用會導致細菌抗藥性菌株的產生及基因污染的疑慮。因此發展非抗生素抗性基因的篩選策略實為當務之急。本計畫預定以一年(1)完成自食用菇釣取同源琥珀酸去氫&#37238;基因及其啟動子基因;(2)進行特定序列的點突變後，建構抗萎銹靈的篩選質體；(3)以農桿菌轉形法建立有效的食用菇菌異源基因轉形系統非抗生素抗性基因的篩選策略；(4)測試其篩選標記基因的穩定性。<br> Abstract: Mushrooms, exhibiting immuomodulatory, anti-aging and anti-tumor activity, are important natural sources of foods and medicines. The combination of traditional mushrooms with the healthcare industry will tremendously increase the value of mushroom products and upgrade the mushroom industry. As the development of the modern biotechnology, the mushrooms provide not only a source of specific genes, but also a bioreactor of specific recombinant proteins. Their function can be further evaluated by cell and animal experiments. By using breeding and hybrid technology, it is possible to select a new strain with specific therapeutic functions for the registration of intellectual properties. The registered strains along with the cultivation techniques can be transferred to the growers. The development of heterologous gene expression in mushrooms will be the breakthrough for the modern biotechnology in the application of molecular pharming. However, the current mushroom transformation systems rely on the selection of antibiotic resistance, such as hygromycin etc. The use of antibiotic resistant marker genes will lead to the occurrence of antibiotic resistant bacteria, as well as the concerns of gene contamination. Therefore, it is urgent to develop a non-antibiotic resistant selection strategy. This project is expected within one year (1) to clone the homologous succinate dehydrogenase gene (sdi1) and its promoter gene, (2) to construct a caroxin resistant selection plasmid by point mutating the specific sequence of sdi1, (3) to establish the heterologous gene expression via Agrobacterium-mediated transformation systems using non-antibiotic resistant selection strategy, and (4) evaluate the stability of selection gene in transformants.異源蛋白質表達分子農場轉基因菇琥珀酸去氫&#37238萎銹靈heterologous protein expressionmolecular pharmingtransgenic mushroomssuccinate dehydrogenasecarboxin