植物防禦素(plant defensins)在各種植物中扮演著抵禦微生物的角色，是由45至54個胺基酸組成的小片段胺基酸鏈。植物防禦素家族中的某些成員，負責平日對於外來各式病原的防禦，一部分成員除了這項工作之外，還與其他受外來病原誘導表現的成員，共同更深入地保護植物免於受到傷害。這樣的誘導表現與茉莉酸甲酯(methyl jasmonic acid, MJ)及乙烯(ethylene, ET)有關。由高氏柴胡(Bupleurum kaoi)的茉莉酸甲酯扣除式資料庫(subtraction library)中，本研究選殖到了兩個防禦素基因，分別命名為BkDF1及BkDF2。根據與其他相關防禦素的序列比對，及純化後的BkDF1蛋白質以蛋白酶酶切處理的結果，推測這兩個防禦素基因隸屬於植物防禦素的第二分群，並且暗示著某種程度的抗微生物特性。為了探討防禦素的特性，本研究在細菌Escherichia coli及酵母菌Pichia pastoris中，分別表現了BkDF1全長及其成熟區域的部分。結果發現表現BkDF1會對E. coli產生毒性。而在酵母菌中所表現出的BkDF1成熟區域蛋白質，則以數種真菌及細菌試驗其抗微生物的特性。除此之外，也分析了BkDF1基因的轉錄程度，結果顯示BkDF1至少負責在柴胡根部及葉片部位對微生物的防禦，且為茉莉酸訊息傳遞路徑中的下游基因。而其表現被水楊酸(salicylic acid, SA)抑制，則印證了植物中兩大抵禦系統，茉莉酸與水楊酸訊息傳遞路徑間的拮抗關係。未來將進一步研究此兩個防禦素基因的啟動子(promoter)區域，以了解更多關於此類基因的調節。

Plant defensins are small peptides of 45 to 54 amino acids undertaking the antimicrobial role in various plants. Part of the family members is responsible for the constitutive guards against pathogen attack. In addition, some members are activated by exogenous agents to further plant protection. This activation is related to methyl jasmonic acid (MJ) and ethylene (ET). From a MJ subtraction library of Bupleurum kaoi, two defensin genes, BkDF1 and BkDF2, were cloned. According to the sequence comparison with other related defensins and protease digestion result of the purified BkDF1 protein, these two genes belong to group II of plant defensins and a considerable antimicrobial activity was implied. To characterize the defensins, BkDF1 and its mature region were expressed in Escherichia coli and Pichia pastoris separately. Expression of BkDF1 is toxic to E. coli. The antimicrobial activity of mature BkDF1 protein expressed in P. pastoris was tested against several fungi and bacteria. In addition, the transcription level of BkDF1 was studied. Our results suggest that BkDF1 may protect roots and leaves against microorganisms and is a downstream gene of the jasmonic acid (JA) signal pathway. Its expression is inhibited by salicylic acid (SA) and demonstrates the antagonistic relationship between JA and SA signal pathways, which are the two major defense response systems in plants. The promoter regions of these two genes will be further investigated to understand more about the regulation of defensin genes.

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