ISSR Analysis Of Germplasm Resources Of Fujian Canavium Album Raeusch

Olive(Canarium album Raeusch) belongs to Burseraceae Canarium L., which originated in China. It is one of the most well-known special fruit trees in south of China, which is distributed in Guangdong, Fujian, Taiwan, Sichuan, Zhejiang, Guangxi and Hainan . Olive has been cultivated in China more than 2000 years. Research on olive mainly focus on cultivation, physiology and medicine composition analysis, etc. But it is seldom that research in molecular biology.Taking 60 Olive genetic resources in fujian as material, this research explored a extraction method for Olive leaf genomic DNA; The Olive ISSR reaction system is established and optimized; 17 primers have been selected which have clear and high polymorphism; Relationship and genetic diversity of fujian Olive genetic resources has been researched by ISSR combined with clustering analysis. The research provided scientific basis for effective protection and rational utilization of Olive germplasm resources. The main results were as follows:1. Facing the fact that it is difficulty to extract DNA from Olive which contains phenolic compound and quite a lot of polysaccharose, the researcher used improved CTAB method to extract DNA of Olive, which meets ISSR requirements in quantity and purity. A set of simple and quick method for olive DNA extraction and purification technology has been found.2. The technological system of ISSR-PCR reaction of Olive has been established. In the 20μL reaction system, the best condition of Olive 1SSR-PCR amplification is 40 ng DNA template, 0.2 mmol/L dNTPs, 0.25μmol/L primer, 1U Taq DNA polymerase, 2.0μL 10×Buffer solution. According to the system, the PCR amplification steps are as follow: 5 min predenaturation at 94℃,30 seconds denaturation at 94℃, 60 seconds annealing, 90 seconds extension at 72℃. The cycle repeats forty times and ends in 10 min extention at 72℃. The system can amplified clear and good effect products, which can meet ISSR analysis.3. 17 ISSR primers were selected from 96 ISSR primers, and 60 Olive germplasm were researched by ISSR method. 151 amplification clips were detected,of which 102 were polymorphic clips, and polymorphic rate is 67.6% . In this research we found that primers containing(TC)n, (CA)n sequence resulted in higher levels of amplified polymorphism of Olive. This is due to the abundant content (AG)n/(GT) n sequence in Olive genome.4. According to results of the clustering analysis to 60 Olive varieties by ISSR.:It proved that the genetic similarity index was between 0.819～0.991. From the result, genetic similarity and relationship among different olive breeds were revealed. It laid theoretical basis for breeding and species introduction on purpose, shortening the breeding period and improving the efficiency of breeding.5. GST of the olive population is 0.2529 .The result showed that olive genetic differentiation mainly exists in the population . According to GST ,the gene flow was estimated Nm=1.4773 ,greater than 1 .It indicated that there exist certain gene exchange between species.