Population Genetics And Speciation Study Of Pinus Armandii And Its Three Affinities

The detailed influence of geographical isolation caused by uplift of mountain and climate instability arised from glacier development on the speciation progress of evolution can be explained by the geographical distribution pattern of genetic variation and differentiation history study. P. armandii is an endemic Pinus plants in the central and western regions of China, which belong to Sect. Cembra of Subgen. Strobus and is the main afforestation tree species and forest regeneration species in China. P. koraiensis, P. griffithii and P. pumila are the closely related species to P. armandii, but there are obvious islation on their living environment and natural geographical distribution area as well as different characteristics on morphology, wood anatomy and cytology.This dissertation studied the genetic diversity and population genetic structure of P. armandii, assessed the gene flow among populations, and then discussed the cause of intraspecific differentiation such as geographic isolation and environment factors, Meanwhile, we also studied the nucleotide polymorphisms and speciation history of P. armandii and its three relatives P. koraiensis, P. griffithii and P. pumila by six nuclear gene loci. The main results were as follows:Our study was based on 696 individuals from 52 natral populations, using 12 SSR markers for PCR amplification and carrying on the genotyping by capillary electrophoresis, then statistical data analysis. Results show that P. armandii has a high level of genetic diversity:He= 0.6045, Ho= 0.4390, (I)= 1.1224, PPL= 0.9167, genetic variation mainly exists between the geography groups (13.67%) and in populations within group (23.82%) and individual in population (58.04%). STRUCTURE cluster and PCA analysis results show that P. armandii populations can be divided into three distinct geographic clusters: populations distributed from the east of sichuan province to the central of China formed a cluster; populations from southwest sichuan and yunnan clustered together and then populations in the eastern Tibet and northwest yunnan was geared to the thired cluster. At the same time, results from the single copy nuclear gene (scnDNA) and chloroplast gene (cpDNA) marker provide further support to the conclusion that P. armandii populations differentiated into two large genetic lineage, although there is no the third cluster of populations in eastern Tibet separated out. In addition, results (Fst) from three nuclear gene loci and two segments of the chloroplast showed strong genetic differentiation between P. armandii populations (Fst value were 0.24548,0.24416,0.22603,0.24416 and 0.22603). We also found that there are high levels of nucleotide polymorphisms in P. armandii (cpDNA:Hs= 0.373, HT= 0.729; scnDNA:Hs= 0.771, HT= 0.872), which may be concerned with the obvious intraspecific genetic differentiation.Data analysis based on SSR, scnDNA and cpDNA markers indicated that geographical isolation events induced by orogenic movement may hinder the geneflow between P. armandii populations and give rise to allopatric speciation of intraspecific spectrum. The result also was endorsed by the significant relationship between genetic distance and geographic distance (r= 0.633, P= 0.633). In the meantime, we detected significant positive correlations between genetic distance and altitude gradient of P. armandii population (r= 0.536, P= 0.01), same situation found between populations differentiation value Fst VS elevation gradient (r= 0.534, P= 0.01) and which revealed that ecological environment factors plays an important role in population genetic division. Furthermore, conjoint analysis results based on microsatellite data of medium-short-term evolutionary history and nuclear and chloroplast genes sequence data of long-term evolution scale demonstrated that geographical isolation and climate change generated by the uplifting of the southeastern edge of the QTP region gave rise to the intraspecific differentiation of P. armandii natural populations, of which the geographic isolation was the main cause. Beyond that the oscillation in Quaternary glaciation and interglaciation and adaptive evolution to different geographic ecological environment may be the deeper reasons of P. armandii intraspecific differentiation. Moreover, study about dynamic history of P. armandii discovered that several populations such as 405、ZD、LP、MN、YY、LS had suffered bottleneck effect. The significantly negative value of Tajima’ D and Fu and Li’s D* and F* based on nuclear genes sequences showed that P. armandii natural populations may have experienced expansion. Species distribution area simulation showed that the suitable area of P. armandii changed little in different periods. The optimum distribution area mainly concentrated in the qinling mountain area in central China and Yunnan-Guizhou Plateau and the east of Tibet, what is more, populations in these regions have higher population genetic diversity and also also focus on the existence of a large number of unique haploid type. So, we speculate that these regions may be glacial refugia for P. armandii.With the gradual uplift of QTP, P. armandii populations in sichuan and yunnan regions were largely affected by the temperature reduction and suffered bottlenecks. While in interglacial, with temperature gradually warming, recent expansion event happened to them.We detected the nucleotide polymorphisms and species differentiation history of four closely related pines Pinus armandii, P. koraiensis, P. griffithii and P. pumila based on sequence data of six nuclear loci. P. pumila showed the highest silent nucleotide diversity (πsil = 0.00464) while P. griffithii had the lowest (πsil= 0.00137), while the value of P. koraiensis and P. armandii falled in between (P. koraiensis:0.00323; P. armandii:0.00269). Analysis of population genetic structure showed that variation primarily existed within populations (74.35%) and there were higher level of genetic differentiation between populations (Fst= 0.576***). which may presumably be due to the habitat fragmentation or the island-like distribution of the Pinus species. Genetic introgression analysis declared that asymmetric gene flow between these four pine species were detected. The effective population size of P. pumila and P. griffithii is smaller than its ancestors’which illustrated that the two species may have experienced bottlenecks. The relationship between P. armandii and P. pumila was closer and gathered as sisters groups while P. koraiensis and P. griffithii clustered in a branch. The four allied species splits into two groups in about 1.37 Mya, and later the differentiation of P. armandii and P. pumila happened in 1.13 Mya. Then P. koraiensis and P. griffithii became divided 970 thousand years ago. The results above support P. armandii is the more ancient specie separated early.