利用ISSR分子標誌分析燈籠草屬物種之遺傳歧異度

Genetic Diversity Analysis of Kalanchoe Species by Inter Simple Sequence Repeat (ISSR) Molecular Markers

本研究利用簡單序列重複區間（inter simple sequence repeat; ISSR）分析蝴蝶之舞、鵝鑾鼻燈籠草綠葉型、鵝鑾鼻燈籠草、掌上珠、大還魂、落地生根及倒吊蓮等7個燈籠草屬物種之遺傳歧異度。由英屬哥倫比亞大學（University of British Columbia; UBC）合成核酸引子的第9組引子中，篩選出多型性佳的19個引子，平均每個引子增幅的片段有10-22條，總共可產生301個增幅的DNA片段，平均每個引子可增幅15.8條，其中多型性條帶有297條，平均每個引子增幅的多型性條帶為15.6條。所有引子的平均多型性比率為98.9%。在19個引子中，在3’端加上核苷酸的引子有17個，僅有2個屬於5’端加上核苷酸的引子。引子以雙核苷酸重複序列的效果較佳，其中以（GA）n及（AG）n重複序列者為多。鵝鑾鼻燈籠草綠葉型、鵝鑾鼻燈籠草、大還魂及倒吊蓮四者相互間之遺傳相似度介於0.576至0.6945之間；其中，以鵝鑾鼻燈籠草綠葉型與鵝鑾鼻燈籠草的遺傳相似度最高，其次為鵝鑾鼻燈籠草綠葉型與倒吊蓮。上述四者與掌上珠、落地生根或蝴蝶之舞之遺傳相似度介於0.150至0.219之間。掌上珠、落地生根及蝴蝶之舞相互間之遺傳相似度在0.195至0.320之間。經ISSR之遺傳相似度分析後，可將燈籠草屬7個物種分為3群。第1群包括鵝鑾鼻燈籠草綠葉型、鵝鑾鼻燈籠草、大還魂及倒吊蓮，第2群為掌上珠及落地生根，蝴蝶之舞則獨立自成一群。 The genetic similarity of seven species of Kalanchoe (Kalanchoe fedtschenkoi, K. garambiensis 'Green Leaf', K. garambiensis, K. poincarei, K. gracilis, K. pinnatum and K. sphaulata) were evaluated according to its inter simple sequence repeat (ISSR). In the 9^(th) kit of University of British Columbia kits, there were 19 primers which induced polymorphic 10-22 marker of each primer. In 19 selected primers, three hundred and one polymorphic bands were generated. On average, there were 15.6 polymorphic bands per primer, showing a very high level of polymorphic (98.9%). The result of ISSR revealed that most of primers, which amplified more markers, contained dinucleotides duplicated sequence. The 3'-anchored primers, poly (GA)n and poly (AG)n motif, were more abundant. The genetic similarity ranged between 0.576 to 0.694 among Kalanchoe garambiensis 'Green Leaf', K. garambiensis, K. sphaulata and K. gracilis. The highest genetic similarity was between K. garambiensis 'Green Leaf' and K. garambiensis, followed by K. garambiensis 'Green Leaf' and K. sphaulata. The genetic similarity of above four specie ranged between 0.150 to 0.219 among and K. poincarei, K. pinnatum and K. fedtschenkoi. While the genetic similarity ranged between 0.195 to 0.320 among K. K. poincarei, K. pinnatum and K. fedtschenkoi. Through ISSR evaluation, 7 species were divided into three groups. The first group comprised of K. garambiensis 'Green Leaf', K. garambiensis, K. sphaulata and K. gracilis, while the second group contained K. poincarei and K. pinnatum. K. fedtschenkoiwas alone in the third group.