低溫逆境對綠豆白化幼苗生長、根尖顯微構造、及細胞膜、液泡膜的影響

Abstract

本論文以兩天大的綠豆白化幼苗?實驗材料，探討經低溫(4.5±0.5℃)處理一天、三天、五天、七天後，一些生理反應的變化，包括：(一)幼苗生長的變化；(二)幼苗之氨基酸和醣類滲出量的變化；(三)幼苗根尖分生組織細胞顯微構造的變化；(四)細胞膜及液泡膜上ATPase活性的變化；(五)細胞膜及液泡膜蛋白質類型的變化。 綠豆白化幼苗在4.5℃的低溫下，生長會完全被抑制；低溫處理後再回到26℃下生長三天，則經低溫處理一天的幼苗，生長會受到延緩；而經低溫處理三天以上的幼苗，幼根已不會再生長，表示根尖生長點已受到破壞。而幼苗氨基酸和醣類的滲出量會隨低溫處理天數的增加而升高，此可以作?泠傷害量化的指標。 觀察根尖分生組織細胞的顯微構造，低溫處理三天的根尖細胞，液泡幾乎全部瓦解，其他胞器亦有明顯被破壞的現象，各種膜系都呈破裂而不連續，此可能由於液泡最先被瓦解後，釋放出內容物破壞了其他細胞構造。 利用二相分佈、蔗糖密度梯度、dextran密度梯度等方法來分離細胞膜和液泡膜，確立了至少有80％ ATPase活性的測得是來自細胞膜或液泡膜上ATPase的反應系統後，比較兩種膜ATPase活性在低溫下的改變，得到兩種膜的ATPase皆會隨低溫處理天數的增加而活性減小，而且細胞膜ATPase活性較液泡膜ATPase活性受低溫影響改變大。而細胞膜和液泡膜蛋白質的電泳分析，都會隨低溫而發生改變，液泡膜蛋白質較細胞膜蛋白質變化多。

Two-day-old etiolated mungbean seedlings (Vigna radiate L.) were used to study the effect of a series of chilling on growth, leakage of amino acids and sugar, ultrastructure of root tip, ATPase activity and proteins of plasma membrane and tonoplast. The growth of seedlings was ihibitted at 4.5℃. When transfered to normal condition 26℃ after chilling treatments, the seedlings would grow more slowly than non-chilling-treated seedlings. The radicles of seedlings which had been treated at 4.5℃ for 3 days had no growth back to 26℃ for 3 days. It indicates that the apical meristem of the root tip was destroyed by chilling, and it was supported by observation with transfer electron microscope. The leakages of amino acids and sugar from mungbean seedlings were enhanced by chilling, indicating the seedlings to be injured by chilling. The ultrastructure of meristem cells of root tips in mungbean seedlings was observed by transfer electron microscope. Remarkable ultrastructural changes were detected within 3 days of chilling treatment. All tonoplast was collapsed, and other organelles and structures were injured apparently. Especially all membranes became broken and discontinuous. Maybe it is because that the early disruption of tonoplast gives rise to the release of inclusion in tonoplast and an alteration of the cytoplasmic environment, consequently, results the degradation of the cells. Plasma membrane and tonoplast vesicles were isolated by two-phase partition, discontinuous sucrose gradient and two-step dextran gradient. The inhibitions of vanadate, azide, nitrate and molybdate on ATP phosphohydrolase (ATPase) activities of plasma membrane, mitochondrion and tonoplast and acid phosphatase activity were used to estabolish the reaction mixture for plasma membrane or tonoplast ATPase assay. It can be obtained 80% activity from plasma membrane ATPase or tonoplast ATPase in estabolished reaction mixture. Plasma membrane and tonoplast ATPase activities were decreased by chilling treatments. And the plasma membrane ATPase was more sensitive to low temperature than the tonoplast ATPase. But the changes of the protein patterns of the plasma membrane caused by chilling treatments were less than those of the tonoplast proteins.