金門稀有植物遺傳多樣性調查
一、研究緣起
金門國家公園為我國第六座國家公園，所處之金門縣於地理位置上和福建省廈門約距10公里，和台灣本島距離約277公里，因此其所分佈的植物種類多與中國大陸相似。於過去所調查的維管束植物中有八屬並無在台灣分佈，此外還有三十五種植物並未見於台灣，其植被具有相當的獨特性。
其中金門地區所產之水韭更為特殊，金門地區之水韭生於太武山區，其生長環境範圍狹小且生態環境變異極大，受當地氣候之影響甚巨；由於金門產水韭棲地狹小，生長區域之水位不穩定，至夏季往往因為過度乾旱而枯死，因此亟需對其生理機制與遺傳特性和其他水韭植物進行比較，以作為金門國家公園未來對金門產水韭棲地保育和管理之策略擬定基礎。
二、研究方法及過程
本研究將以建立水韭屬植物小分子熱休克蛋白之分子標誌，配合RAPD（random amplified polymorphic DNA）方法，並分析水韭屬植物之基因序列等方法，比較金門地區水韭與鄰近區域產水韭之遺傳變異及親緣關係，以釐清其分類地位；並萃取金門地區水韭植物體中RNA，試驗觀察其小分子熱休克蛋白之表現，探討其於該棲地之生長作用機制；同時進行孢子萌發試驗，試驗於不同溫度環境下對其萌芽之影響，以做為日後金門產水韭之保育以及金門國家公園管理之參考。
三、重要發現
在分類地位方面：以聚合?連鎖反應擴增台灣水韭、金門地區水韭之小分子熱休克蛋白sHSP21基因保守序列部分片段，其長度為410 bp，結果發現金門地區水韭與台灣水韭兩者基因序列相似度達96.7％，與寬葉水韭相似度則為90.9％；但以RAPD方法分析所得結果，金門地區水韭與台灣水韭之相似度僅31.42％，與寬葉水韭相似度為34.32％。此結果為首次以嚴謹分子證據，闡釋金門水韭應與台灣水韭有所差異。在復育研究方面：以 5℃處理過之金門地區水韭孢子，其萌發速率較其他溫度處理快，但台灣水韭則是以35℃處理過之孢子有最高之萌發率。
綜合以上各點，可發現金門地區水韭與台灣水韭在分子序列及生理表現上均確實有差異。也因此，前人建議以金門水韭(Isoetes kinmenensis)命名，以區別台灣水韭確實可考慮。而針對金門地區水韭所做之小分子熱休克蛋白sHSP21 mRNA表現量測定實驗，更可做為金門水韭能夠適應金門狹小且嚴苛的生長棲地之理論。
Abstract
Kinmen National Park located off the southeastern coast of Fujian Province in Xiamen Bay at the outlet to the Jiulong River. Due to the close proximity of Kinmen to mainland China, Kinmen's flora is closely related to mainland China's. There are 8 genus vascular plants and 35 species of plants have not found in Taiwan. Among these distinctive plants, the quillwort in Kinmen is more special. This quillwort is grown on Tai-Wu Mountain. The growing condition compared with Taiwan’s is not well owing to the huge dynamic variation of weather and it’s nature vegetation. The habitat of the quillwort in Kinmen is Narrow and small. In summer, the water level of the growth region is unstable leading to the death of quillwort. Therefore, based on the concern for wildlife conservation and the natural environment, We need to investigate and compare the physiological mechanism and the hereditary property of the quillwort in Kinmen with other species.
The aim of the study is to establish the specific molecular sequence and analyzing the sequence of small heat shock protein. In this way we can compare the hereditary variations and reconstruct phylogeny of Isoetes. Furthermore, optimal spore germination will be studied.
By amplifying the sHSP21 partial conserved sequence of I. taiwanensis and the quillwort in Kinmen with Polymerase Chain Reaction, we found sequence variation among these two quillworts. Besides, this primer could amplify a unique sequence in the quillwort in Kinmen which is not found in I. taiwanensis. The RAPD analysis also showed only 31.42% similarity between them. In addition, the spore germination as well as temperature response varies between these two samples. All of the data of this research indicated that a new species called Isoetes kinmenensis might be the right name for Kinmen quillworts.
金門國家公園為我國第六座國家公園，所處之金門縣於地理位置上和福建省廈門約距10公里，和台灣本島距離約277公里，因此其所分佈的植物種類多與中國大陸相似。於過去所調查的維管束植物中有八屬並無在台灣分佈，此外還有三十五種植物並未見於台灣，其植被具有相當的獨特性。
其中金門地區所產之水韭更為特殊，金門地區之水韭生於太武山區，其生長環境範圍狹小且生態環境變異極大，受當地氣候之影響甚巨；由於金門產水韭棲地狹小，生長區域之水位不穩定，至夏季往往因為過度乾旱而枯死，因此亟需對其生理機制與遺傳特性和其他水韭植物進行比較，以作為金門國家公園未來對金門產水韭棲地保育和管理之策略擬定基礎。
二、研究方法及過程
本研究將以建立水韭屬植物小分子熱休克蛋白之分子標誌，配合RAPD（random amplified polymorphic DNA）方法，並分析水韭屬植物之基因序列等方法，比較金門地區水韭與鄰近區域產水韭之遺傳變異及親緣關係，以釐清其分類地位；並萃取金門地區水韭植物體中RNA，試驗觀察其小分子熱休克蛋白之表現，探討其於該棲地之生長作用機制；同時進行孢子萌發試驗，試驗於不同溫度環境下對其萌芽之影響，以做為日後金門產水韭之保育以及金門國家公園管理之參考。
三、重要發現
在分類地位方面：以聚合?連鎖反應擴增台灣水韭、金門地區水韭之小分子熱休克蛋白sHSP21基因保守序列部分片段，其長度為410 bp，結果發現金門地區水韭與台灣水韭兩者基因序列相似度達96.7％，與寬葉水韭相似度則為90.9％；但以RAPD方法分析所得結果，金門地區水韭與台灣水韭之相似度僅31.42％，與寬葉水韭相似度為34.32％。此結果為首次以嚴謹分子證據，闡釋金門水韭應與台灣水韭有所差異。在復育研究方面：以 5℃處理過之金門地區水韭孢子，其萌發速率較其他溫度處理快，但台灣水韭則是以35℃處理過之孢子有最高之萌發率。
綜合以上各點，可發現金門地區水韭與台灣水韭在分子序列及生理表現上均確實有差異。也因此，前人建議以金門水韭(Isoetes kinmenensis)命名，以區別台灣水韭確實可考慮。而針對金門地區水韭所做之小分子熱休克蛋白sHSP21 mRNA表現量測定實驗，更可做為金門水韭能夠適應金門狹小且嚴苛的生長棲地之理論。
Abstract
Kinmen National Park located off the southeastern coast of Fujian Province in Xiamen Bay at the outlet to the Jiulong River. Due to the close proximity of Kinmen to mainland China, Kinmen's flora is closely related to mainland China's. There are 8 genus vascular plants and 35 species of plants have not found in Taiwan. Among these distinctive plants, the quillwort in Kinmen is more special. This quillwort is grown on Tai-Wu Mountain. The growing condition compared with Taiwan’s is not well owing to the huge dynamic variation of weather and it’s nature vegetation. The habitat of the quillwort in Kinmen is Narrow and small. In summer, the water level of the growth region is unstable leading to the death of quillwort. Therefore, based on the concern for wildlife conservation and the natural environment, We need to investigate and compare the physiological mechanism and the hereditary property of the quillwort in Kinmen with other species.
The aim of the study is to establish the specific molecular sequence and analyzing the sequence of small heat shock protein. In this way we can compare the hereditary variations and reconstruct phylogeny of Isoetes. Furthermore, optimal spore germination will be studied.
By amplifying the sHSP21 partial conserved sequence of I. taiwanensis and the quillwort in Kinmen with Polymerase Chain Reaction, we found sequence variation among these two quillworts. Besides, this primer could amplify a unique sequence in the quillwort in Kinmen which is not found in I. taiwanensis. The RAPD analysis also showed only 31.42% similarity between them. In addition, the spore germination as well as temperature response varies between these two samples. All of the data of this research indicated that a new species called Isoetes kinmenensis might be the right name for Kinmen quillworts.