Spectral Geography And Conservation Genetics Of Rare Dendrobium

Dendrobium offcinale Kimura et Migo (Orchidaceae) is an endemic and endangered herb distributed in China. Its stems have been used as a traditional Chinese tonic medicine named "Tiepi Fengdou" for hundreds of years, which benefits people in many ways according to the record in "Ben Cao Gang Mu" and "Pharmacopoeia of the People’s Republic of China". Its main function is to nourish kidney and clear away heat-evil, benefit the stomach and promote the production of body fluid,moisten the lung and relieve cough, resist cancer and prolong life. The limited natural resource and great demands result in the over-exploitation of its germplasm resources,which in turn incents the depredation of this species again as a vicious circle. It is so urgent for us to study its phylogeography and conservation genetics.For understand the delimitation of Dendrobium sections and investigate the evolutionary status of D. officinale, we undertook a comparative study between morphological and molecular phylogeny at section-level of Dendrobium. Maximum Parsimony approach was .used to analyze 38 morphological characters among 16 sections. Molecular data of 405 nrDNA ITS ribotypes were obtained from 740 accessions of 160 species that were analyzed by Maximum likelihood, Bayesian inference and Bayesian skyline analyses. The results showed that nuclear phylogeny was mostly congruent with morphological taxonomy. Based on morphological characters and molecular data, 12 sections within 16 sections could be recognized as four section complexes: Formosae complex, Crumenata/Aporum complex Calcarifera/Pedilonum complex, and Dedrobiun/Holochrysa complex. Section Grastidium and Stachyobium were confirmed to be monophyletic with strong support.The species of sect. Densiflora were embedded into nearly all clades of Asian clade,its evolution position was complicated that needed more sampling for reevaluating.Similar to the Dendrobium section complexes, species complexes also existed in Dendrobium genus. For 160 species, 54 species were evaluated as monophyletic species and 6 species complexes were identified. Excluding D. moniliforme complex,other five species complexes were first identified in Asian Dendrobium,containing D.officinale complex, D. nobile complex, D. densiflorum complex, D. strongylanthum complex and D. fimbriatum complex. Our results indicated that it was preferable to establish section complexes of Dendrobium for evaluating the evolution of Dendrobium genus. For D. officinale complex, D. tosaense, D. scoriarum, and D.flexicaule could be considered as D. officinale complex or closely relative taxa.In this study, some molecular techniques including nrDNA ITS and non-coding region of cpDNA will be used for defining haplotypes and ribotypes of D. officinale and closely relative taxa, understanding its geographic distribution and reconstructing its haplotypes and ribotypes phylogenetic trees. Compared with outgroups, the ancient haplotype and its distribution area will be defined. The incidents that ever happened to different clades can be deduced using network analysis method. The results showed that D. Tosaense and D. officinale were not genetically distinct from one another.Rapid radiation and adaptive evolution probably drove speciation of D. flexicaule.Discordances between chloroplast and nuclear genes were also found: potential gene introgression was found between D. officinale and D. scoriarum; Incomplete lineage sorting, hybridization or introgression form integral aspects of adaptive radiation probably drove speciation of D. shixingense. Gene flow via pollen in D. officinale is more than that via seeds. Current highly fragmented and discontinuous distributions of D. officinale were due to recently over-exploitation.We also developed a set of polymorphic nSSR markers for D. officinale and described the genetic connectivity of natural populations of D. officinale throughout China. Included comprehensive population-level sampling of D. officinale, aim of this project was to survey genetic diversity, detect population structure, and determine if recent habitat loss and fragmentation have caused genetic bottlenecks. Based on phylogeography and conservation genetics of D. officinale, it is helpful for the determination of the ESUs and the conservation strategies of D. ofcinale. For conservation management plans, four evolutionary significant units of D. officinale were treated as separate management units for which in situ conservation, including habitat restoration, should ensure persistence of this species and preservation of its evolutionary potential.To evaluate the effectiveness of ex situ collection of D. officinale genetic diversity,we developed 15 polymorphic trinucleotide microsatellite loci of D. officinale to examine the genetic diversity and structure of three D. officinale germplasm collections and their respective wild populations in China. The three germplasm collections showed reductions in gene diversity and average number of alleles per locus, but an increase in average number of rare alleles per locus in comparison to their wild populations. However, the differences in gene diversity between the germplasm collections and wild populations were not statistically significant. The analysis using STRUCTURE revealed evident differences in genetic composition between each germplasm collection and its wild population, probably because the D.officinale individuals with distinct genotypes in each wild population were unevenly selected for establishing its germplasm collection. For conservation management plans, we propose that D. officinale individuals with rare alleles need to be conserved with top priority, and those individuals with the most common alleles also should be concerned. The 15 new microsatellite loci may be used as a powerful tool for further evaluation and conservation of the genetic diversity of D. officinale germplasm resources.