The Study On The Genetic Diversity Of Mytilaria Laosensis By ISSR

Mytilaria laosensis is an evergreen tree of hamamelidaceae mytilaria, which is one of the important native tree species in Guangxi. Mytilaria laosensis not only is a good ornamental tree with straight trunk, thinner branches and elegant burlywood, but also has high economic and ecological values. It has been listed as the third-level protected plant by the province. Mytilaria laosensis, which is a sensitive plant to climate, widely distribute in tropical and southern subtropical regions, and now has been successfully introduced to Hunan province and other places. In this study, the ISSR markers was used to analyse the genetic diversity of mytilaria laosensis resources, which could determine the living conditions of mytilaria laosensis populations from the molecular level. At the same time, the trend of the distribution of natural populations of Mytilaria laosensis was speculated by Analysis and simulation of meteorological data. This is scientific foundation and guidance for its Northward movement.Based on the material of6populations with69samples, through the single factor design, the various factors affecting ISSR-PCR reaction were studied to determine the optimum concentration range for each factor, then through orthogonal design and variance analysis, the optimal system of ISSR-PCR reaction was established as follows:30ng template DNA,0.20mmol/L dNTPs,2.75mmol/L Mg2+,0.6μmol/L primer,2.20U Taq DNA polymerase were contained in20μL reaction system. Variance analysis showed that: only the Mg2+concentration of five factors has the greatest impact on the reaction system and reached a significant level.According to the optimal ISSR-PCR reaction system, through the combination of primer screening and the determination of annealing temperature,9primers screened from100random primers, which have high polymorphism and good repeatability, were applied to ISSR-PCR amplification of Mytilaria laosensis samples. Total129DNA bands were obtained, of which were109polymorphic bands. The range of bands length augmented by each primer was between180to2000bp, and the percentage of polymorphic bands was84.50%. It showed that the rich genetic diversity was existed in the mytilaria laosensis resources. The highest genetic diversity of the six groups is Pingxiang and the minimum of the genetic diversity is the Napo.The data analyzed by the PopGen32software revealed that the polymorphic loci (PPB) percentage of mytilaria laosensis was84.50%; gene diversity(H) index was0.2969; Shannon diversity index(I) was0.4404. The result showed that mytilaria laosensis has a high genetic diversity at the species level. This genetic diversity was much higher than other endangered species, but slightly lower than the widespread species, which illustrated that it is related to its distribution and breeding methods. Genetic diversity (Ht) of mytilaria laosensis groups was0.3032; genetic diversity within the group was0.2512; coefficient of gene differentiation(Gst) was0.1716; variation within species accounted for82.85%, variation among species accounted for17.15%. The above information indicated genetic differentiation among6mytilaria laosensis populations is relatively small, and the diverse microhabitat is the main reason for population differentiation. The gene flow(Nm) among populations was2.4134, which meaned that there exists certain gene flow in the group, and it is much enough to resist the influence of genetic drift.On the basisi of the genetic consistence and Nei's (1978) genetic distance, the6populations were analyzed by UPGMA cluster analysis, which displayed that Hunan Zhuzhou population and Hunan Xining one had the closest genetic relationship and the relationships between Guangxi Pingxiang population and the above two Hunan province ones were closer than the remaining other tested groups. This also proves their introduction relationship, Zhuzhou groups were introduced to Hunan Xinniing and Hunan Xinning group were introduced to Guangxi Pingxiang.As to the introduction relationship, Hunan groups and Guangxi Pingxiang groups had shown little difference not only in genetic polymorphism but also in the index of genetic diversity. Meanwhile, it also demonstrated that plants native to tropical and southern subtropical regions could grow normally in Hunan Zhuzhou and Hunan Xinning. This provides a strong evidence for global warming, and also provides some theoretical support for the mytilaria laosensis population northward movement.According to the result of the RegCM3meteorological model, natural populations of mytilaria laosensis may be distributed to about25°N and extends to about26°N in coastal zone in2042.