五節芒族群對鉛忍受力之生化學探討

Abstract

五節芒（Miscanthus floridulus）在臺灣為廣泛分佈種，其族群遍佈於臺灣各地，包含最惡劣的環境如裸地、火山口、重金屬污染和重金屬礦區等地，其適應性為本省草原植物之冠。本研究的主要目地係為瞭解鉛污染地區（高速公路42公里）與非鉛污染地區（南港中央研究院）五節芒植物族群在對於抗鉛的能力上，是否因為環境中鉛的含量不同，而產生不同的生態型（ecotype），此生態型，是否因環境篩選而產生遺傳性變異等問題加以探討。 在探討鉛污染與非鉛污染地區的五節芒植物族群的抗鉛性研究，以生物分析及水耕的方式，測定不同地區的五節芒族群對重金屬鉛的忍受能力，其結果顯示五節芒族群在鉛污染地區生長者有較高的抗鉛能力。 在不同鉛濃度溶液下，五節芒種子發芽及生長情形得知，在鉛污染與非鉛污染地區所採得的五節芒種子的發芽率不受鉛濃度的影響，但就初生根和初生莖而言，則鉛污染地區的幼苗生長較好，即鉛污染的地區五節芒族群比未受污染地區的族群具有較高的忍受能力。在幼苗體內的超氧岐化?活性的變化情形，結果顯示鉛污染地區五節芒族群比未受污染地區的族群幼苗體內超氧岐化?活性上升幅度大。由其生長情形與超氧岐化?活性變化情形而言，鉛污染地區的五節芒族群確有生理上的變化而能適應有鉛存在的環境，因而產生出具抗鉛性的族群。 以水耕法，將鉛污染地區與非鉛污染地區的五節芒種子在無鉛環境下發芽生長30天，再以不同濃度的鉛溶液處理20天和70天，比較二族群間植株的生長情形，結果顯示與上述五節芒鉛處理下發芽生長的幼苗頗相似，即在鉛污染地區的五節芒族群的生長情形較好。在鉛處理20天的五節芒植株葉片中，超氧岐化?活性變化不大。在鉛處理70天的植株根部，超氧岐化?活性皆下降，而在鉛污染地區五節芒族群比未受污染地區的族群內超氧岐化?下降幅度較小。在葉部的超氧岐化?活性皆上升，在鉛污染地區五節芒族群葉部超氧岐化?活性上有較大的上升幅度，即在鉛污染地區的五節芒族群確有特殊抗鉛性的出現。 為瞭解此抗鉛性的生態型在遺傳上是否因環境壓力產生變化，以聚丙烯醯胺膠體電泳法進行同功?分析，及和以隨機擴增多型DNA分析方法，計算出族群間的Eclidean離和平均距離，以此數據矩陣進行群叢分析（cluster analysis），結果顯示此具有抗鉛性的五節芒族群在同功?和隨機擴增多型性DNA所得的相似度上，無法將二個重金屬污染地區（高速公路42公里和金瓜石）和四個非重金屬污染地區（南港中研院、南投縣大坪頂、南投縣神木村和蘭嶼）的五節芒族群區分開來。若經長期鉛污染的誘導，或可產生出抗鉛性的生態型。

Miscanthus floridulus is the most dominant native grass in Taiwan. Its population is widely distributed from low to high elevations and can survive the most severe environments, including arid areas, volcanic craters, heavy metal-polluted areas, and heavy metal mines. This study was designed to determine the lead tolerance of M. floridulus populations in lead polluted and non-polluted areas, whether the difference in lead concentration leads to the formation of different ecotypes, and whether such selection pressure leads to genetic divergence. To determine the lead tolerance of M. floridulus in lead polluted and non-polluted areas, bioassay and water culture were used. The results showed that the populations in lead polluted area have higher lead tolerance. In the bioassay, the seeds of Miscanthus floridulus collected from lead polluted and non-polluted areas were moistened with a series of concentrations of lead aqueous solutions, 20, 40, 50, 60, 80, 100, 200, 300 and 500 ppm. Results showed that, seed germination rates of both populations were not affected by lead concentration. Inhibition of seedling growth increased with the rise in lead concentration. Such inhibition was less prominent in the population from lead polluted areas, suggesting that this population have higher lead tolerance than the one from non-lead-polluted areas. The results from seedling SOD activity suggested that the population from lead polluted areas showed higher increase in SOD activity. The results from the bioassay suggest that the population from lead polluted areas are physiologically adapted to lead stress, and have developed into a lead tolerant ecotype. In the water culture experiment, seeds were first put in lead-free Kimura solution to germinate and grow for 30 days, then treated with different lead concentrations (50, 100, 150, and 200 ppm) for 20 and 70 days. The results were similar to those of the bioassay tests, which indicate that the population from lead polluted areas are less inhibited by lead than those from non-lead-polluted areas. The leaf SOD activity tested after 20 days showed no significant change. The root SOD activity tested after 70 days decreased with-increased lead concentration, and such decrease was less prominent in the population from lead polluted areas. The leaf SOD activity tested after 70 days increased with increased lead concentration, and such increase was more significant in the population from lead polluted areas than from non-polluted areas, suggesting lead tolerance in lead polluted area population. To further understand if the lead tolerant ecotype had changed at the genetic level, the PAGE and RAPD methods were used to analyze the Euclidean distance and the average distance between the populations of M. floridulus. Cluster analysis was used to produce dendrograms of the populations. The results from population similarity could not distinguish the two populations at heavy-metal polluted areas from the four populations grown at non-heavy metal polluted areas. This result suggests that the lead tolerant ecotype of M. floridulus from lead polluted areas are not different genetically, but induced by lead pollution.