惠蓀實驗林場關刀溪流域靈芝屬與烏芝屬之分類研究

Abstract

蕙蓀林場關刀溪流域為全球長期生態調查研究計畫中的一站，我們針對蕙 蓀林場中關刀溪流域原始林進行調查，分離出14株靈芝屬及烏芝屬真菌， 依外型可分為三群。以逢機增殖多型性核酸(RAPD)及核糖核酸基因非轉錄 區(rDNA ITS1)定序實驗，進行分類研究。測試的20條RAPD 引子(primer) ，其中有16條引子具有多型性產生。我們使用七條引子共計獲得145個不 同的條帶(band)，將所得的電泳圖記錄為二元資料，以RAPDistance軟體 計算各菌種之間的相似度，採用UPGMA程序進行歸群，以NTSYS-PC軟體進 行矩陣相關性比較、主座標軸分析及最小展開樹，亦可歸為三群。在DNA 定序方面，使用PCR合成雙股rDNA ITS1區域約290 bp的片段，進行PCR產 物直接定序，DNA序列以Clustal W軟體進行最相似排列，並以Kimura''s 2-parameter方法計算DNA序列之間的核酸距離，並加以歸群，以GCG序列 分析軟體比較各序列之間相似度，並與Genbank所取得的DNA序列比對，可 鑑定出其接近的種類。DNA序列分析支持RAPD實驗所得的歸群結果。

Huisun Experimental Forest is a station of LTER ( Long Term Ecological Research ). Isolates of Ganoderma spp. from Huisun can be characterized into tree races by means of biological typing. DNA from 14 isolates was amplified by the random amplified polymorphic DNA (RAPD) technique by using the polymerase chain reaction with single primers. The amplification products were analyzed for polymorphisms by gel electrophoresis to determine whether species could be distinguished at the molecular level. The amount of genetic variation was evaluated by employing polymerase chain reaction amplification with a set of 7 random 10-mer primers. All amplifications revealed scorable polymorphisms among the isolates, and a total of 145 band positions was scored with binary coding for the 7 primers tested. UPGMA (unweighted pair-group method using arithmetic averages) cluster analysis and PCOA (principal coordinate analysis) were used to divide the Ganoderma sp. isolates into three distinct clusters.Genetic distances between each of the isolates were calculated by RAPDistance software, and cluster analysis was used to generate a dendrogram showing relationships between them. The results indicate that RAPD markers can be a quick and reliable alternative for differentiating isolates of Ganoderma spp. rDNA ITS-1 was amplified by polymerase chain reaction(PCR) the products were used for sequencing. ITS-1 data for Ganoderma are generally consistent with the result of RAPD.