Molecular Phylogeographic And Genetic Diversity Of Taxus L. (Taxaeae) In China

Taxus has plenty of resources wide distribution in China, the Flora of China recorded 3 species and 2 varieties of this genus(Taxus cuspidate, T. wallichiana, T. fuana, T. wallichiana var. mairei, T. wallichiana var. chinensis)(1999). but its wild trees have often been severely endangered due to over exploitation of the resources. Therefore, understanding genetic diversity and structure of natural populations of Taxus is of significant theoretical importance for evaluation and conservation of its genetic resources. Molecular phylogeographic studies would provide strong evidences for understanding the genetic diversity and evolutionary history of Taxus in China. This study collected samples range-wide of almost all species of Taxus in China for molecular analyses, and new microsatellite markers were developed and characterized by screening a number of SSR primers for analyses of genetic diversity and structure of different populations of Taxus. Additional sequence information on 3 chloroplast genes, 1 mitochondrion gene and 1 nucleus gene were used to detect phylogenetic geographical relationships among and genetic diversity of the 5 species(including varieties) of Taxus, the population demographic history of Taxus in China can therefore be determined. The main results were as the followings:(1) We detected a high level of genetic variation across the 64 populations of Taxus from mainland China. 44 different cpDNA haplotypes were detected. Hd=0.7368, π=0.00396 and HT=0.758 were detected in the combined petA-psbE/ rpl16 /rps16 chloroplast region. 6 different mtDNA mitotypes with Hd=0.466, π=0.66×10-3 and HT=0.451 were detected in nad7/3-3 mitochondrial region. 73 different nrDNA haplotypes with Hd=0.9143, π=0.00891 and HT=0.813 were detected in ITS Nuclear gene sequence. Genetic diversity analysis based on 12 SSR polymorphic loci on 786 samples from 41 natural populations showed that 288 alleles(19.244 on average), He=0.479 and Ht=0.498> Hs=0.282. Meanwhile, it was found that populations in Hengduan mountains in southwest China and Qinling-Daba mountains displayed a higher genetic diversity, while populations in east China, central China and northeast Changbai mountains displayed a lower genetic diversity.(2) AMOVA analysis based on cpDNA and mtDNA data showed that the variance among populations was much higher than that within populations, however, analysis based on nuclear gene information(nrDNA sequence and nrSSR) showed that the variance within populations was higher than that among populations. In Taxus, both chloroplast DNA and mitochondrial DNA are of paternal inheritance, allowing an independent assessment of gene flow from paternally plastid DNA via pollen. The unique structure of pollen without air chamber and the occurrence as understory trees, make the dispersal distance of Taxus pollen limited. But nuclear DNA is of biparental inheritance, representing seed flow. At the seed maturity stage, the red aril with sweet taste of seeds would attract birds to eat and spread the seeds. This process facilitates seed flow among populations, therefore reducing differentiation among populations.(3) Strong phylogeographic structure among the populations of Taxus was detected, and the NST was significantly greater than the GST for both nuclear and chloroplast DNA(nrDNA:GST=0.412<NST=0.792;cpDNA:GST=0.7840,NST=0.8280), this is largely due to geographic isolation. According to haplotype strict consensus trees and haplotype network of the cpDNA or nrDNA over all the populations in the study, it is appropriate for haplotypes from the same region to be clustered together. The Mantel test also showed that significant correlation between genetic distance and geographic distance.(4) Detection of historical expansion of populations showed that the Tajima’s D test displayed a neutral mutation evolution, and the mismatching analysis showed no population expansion in recent history, but only analysis of Changbai mountain showed a unimodal curve, suggesting that the population might have experienced expansion. The haplotypes obtained in this study were in two types of distribution: 1) many populations had its unique haplotypes; 2) Some populations shared same haplotype. This phenomenon probably implies that the distribution of Taxus experienced exotic fragmentation and indicates there were multiple refugia for Taxus.