|其他摘要||The evolutionary history of plants in southeast Tibetan Plateau might be the most complicated around the world because of the extremely complex topography and climate in this area induced by its uplift. High biodiversity, rich endemic plant and rapid population declines have led this area and adjacent region become a world biodiversity hotspot. However, so far the studies using molecular markers at the intra-specific level are lacking for plant taxa inhabiting the Tibetan Plateau area and adjacent areas. Dipentodon sinicus is the endemic archaic tree of Eastern Asia restricted to this area, which is listed as vulnerable species in the China Species Red List. We implemented a phylogeography and conservation genetics study using chloroplast sequences on Dipentodon sinicus with an attempt to test how tectonic activities shaped the plant population structure in this area. The results of this study will shed additional light on the evolution of biodiversity on Tibetan Plateau and adjacent areas and provide historical background for the conservation of biodiversity in this area, undoubtedly including D. sinicus.
After investigating pollination mechanism, seed dispersal mechanism and chromosome numbers of D. sinicus, this study revealed genetic diversity and population genetic structure of this species based on chloroplast sequence, discussed how tectonic activities and complex climate in southeast Tibetan Plateau shaped the plant population structure, defined its evolutionarily significant units and suggested the conservation strategy, constructed microsatellite DNA library of D. sinicus for nuclear phylogeography and conservation genetics in next work. The main results are summarized as follows:
1. Reproduction ecology and cytology
The results of anthecology in YIL population showed that D. sinicus was almost completely pollinated by insects, just a little by wind. The main insects recorded as flower-visitors were Eristalis tenax Linnaeus, Eristalis cerealis Fabricius, Episyrphus balteatus Geer., Syrphus vitripennis Meigen, Apis cerana Fabricius and Bombus parthenius Richards. The field observation for the ripe fruits indicated that this species dispersed its seeds through ejecting from capsules. The study of cytology showed that chromosome numbers of individual in YIL and YL populations are all 2n=34.
Based on psbA–trnH and rps16–trnQ sequences data in 16 populations of D. sinicus, the study of phylogeography revealed that D. sinicus had high genetic diversity (hT=0.902) and high genetic differentiation (NST=0.987 and GST=0.948). The high genetic diversity may be attributed to its long evolutionary history and highly diverse habitats. The ineffective mode of seed dispersal and dramatic tectonic elevation in the distribution range of D. sinicus could conspire to cause the high genetic structure in D. sinicus. Nested clade analysis supported that allopatric fragmentation induced by orogenesis could explain the highly differentiated structure. However, the intraspecific relationships of 3-step clades were not consistent with the morphology distinction of D. sinicus. And the haplotype network did not reflect the geographical distribution of 2-step clades, as geographically approximate southeast populations (clade 2-1) and northwest populations (clade 2-2) in Yungui Plateau were the most strongly differentiated. We supposed that a fragmentation of an ancestral population caused by the uplift of Tibet and Yungui Plateaus, resulting in lineage sorting of ancient clades and a loss of alleles through drift may be the function of this pattern. Tectonic movement of northwest Yunnan might be an alternative reason for this phylogeographic pattern. The results of this study demonstrated that significant increases in geological and ecological diversity that accompanied the uplift of Tibet Plateau and adjacent areas have very important impacts on the evolution of biodiversity in this area.
3. Evolutionarily significant units and conservation strategy
For conservation management of D. sinicus, in situ strategy should be emphasized. Ex situ to enhance diversity through transplant or reintroduction from each other is not advisable, in order to avoid loss of critical adaptive features and the possibility of compromised long-term viability due to outbreeding depression. Based on most parsimonious and UPGMA trees, the geographical distribution of genetic variation, genetic distance and AMOVA, four ESUs (ESUA-D) were designated. The gradation of conservation priority is ESU B, ESU A, ESU D and ESU C. Ancestral ESU B (GS,FG1,FG2) should be given the highest priority to its conservation. The second priority should be given ESU A (YJ,YIL,AL) because cpDNA variation in the ESU was highest structured, its three populations must be separately managed in situ. For the most strongly differentiated ESU D (VN,MLP,HS,LS,CJ,TL,RS), it should gotten the third priority, population VN, HS and LS is emphases in the ESU. Finally, the last divergent ESU C (TC,YL,LUS) would be of the lowest conservation priority.
4. Microsatellite DNA library
The genomic DNA was converted into the fragments and hybridized to biotinylated microsatellite oligonucleotides and captured on streptavidin-coated paramagnetic beads (Dynal). Unwanted DNA was washed away and captured’ DNA was cloned and sequenced, and then the largely enriched fragments containing SSR were obtained. The result showed that 91.67% (88) sequences containing the repeat motifs was found in 96 clones, 34 sequences selected from them composed microsatellite DNA library of D. sinicus. This library would establish a foundation for nuclear phylogeography and conservation genetics of D. sinicus in the future.|