Studies of population genetics on four mangrove tree species in Okinawa, Japan

本研究试图分析四种红树树种木本植物木本植物的遗传变异。Kandelia Candel，Rhizophoraceae，Rhizophora stilosa，Sonneratiaceac)，增加了菊科Lumnitzera racemosa一种用于DNA分析。在实验前，从四个树种的树木上采集了树叶作为样本，并将其储存在冷冻箱中。采用垂直平板聚丙烯酰胺凝胶电泳法(PAGE)对4种红树植物的同工酶进行了分析。电泳法检测到23种酶。柱花根霉产5种酶，木霉产5种酶。木本植物只有过氧化物酶(POD)具有多态性，而白海桑(Sonneratia Alba)只有黄递酶(DIA)、酯酶(EST)和葡萄糖激活酶(GR)。DNA提取采用CTAB(十六烷基三甲基溴化铵)法和冰冻缓冲法。冰冻缓冲法提取DNA的效果优于CTAB法。为了获得线粒体DNA片段，使用了18种限制性内切酶。用23个引物扩增叶绿体DNA。对有丝分裂原和基因组DNA片段进行扩增和分析。由于在冲绳采集的所研究的红树林物种的遗传变异很小，所使用的引物并不是那么有效。然而，我们可以获得证据表明，微卫星是分析琉球群岛海鞘根霉遗传变异的有效手段之一。

In this studies, attempts have been made to analyze genetical variation of four mangrove tree species, Bruguiera gymnorrhiza (Rhizophoraceae). Kandelia candel (Rhizophoraceae), Rhizophora stylosa (Rhizophoraceae) and Sonneratia alba (Sonneratiaceac), distributed in Okinawa, Japan, and adding one species, Lumnitzera racemosa (Combretaceae) for DNA analysis. Leaves as samples were collected from trees of four species, and stocked in a deep-freezer before experiment. Polyacrylamide gel electrophoresis (PAGE) of a vertical slab-format method was used for analyzing isozyme loci of four mangroves. After the electrophoresis, 23 enzyme species were stained. Fine zymogrames were obtained five enzyme species for Rhizophora stylosa and five enzyme species for Bruguiera gymnorrhiza. Polymorphism was found only peroxidase (POD) for Bruguiera gymnorrhiza, but diaphorase (DIA), esterase (EST) and glucokinase GR) for Sonneratia alba.DNA was extracted from leaves of Rhizophora stylosa. Both CTAB (Cetyl Trimethyl Ammmonia Bromide) and Ice Cold Buffer methods were applied for DNA extraction. The Ice Cold Buffer method was better than the CTAB method for DNA extraction. To obtain mitochondrial DNA fragments, 18 restriction enzyme species were applied. For amplification of chloroplast DNA, 23 primers were used. Mitokondorial and genom DNA fragments were amplified and analyzed. Since genetical variations were small in studied mangrove species collected in Okinawa, primers applied were not so efficiently worked. However, we could obtain evidence that micro-satellites are one of efficient ways for gentical variation analysis of Rhizophora stylosa in Ryukyu islands.