| The genus Stipa Linn. is placed in the family Poaceae and belongs to tribe Stipeae, but lacking of clear morphological boundaries among taxa. Stipa species distribute throughout a wide variety of habitats existing geographical substitution at a large spatial scale and habitat substitution at small scale. However, it is difficult to reveal the system evolution and regional differentiation based on the traditional morphological analysis. DNA sequences and molecular markers are the effective methods to study the evolution and differentiation of species. In this paper, three gene sequences (rbcL, trnL-F, ITS) were used to analyze the circumscription, the phylogenetic relationships and regional differentiation of the genus taxa collected from Mongolian Plateau and Qinghai-Tibet Plateau and Loess Plateau. The main results are as follows.1. MP (Maximum-Parsimony) trees of Stipa L. from rbcL, trnL-F sequences were consistent. The results revealed that species of Stipa were divided into two major clades. One clade was corresponded with Sect. Leiostipa, Sect. Stipa, Sect. Barbatae and Sect. Smironvia, while the other clade exactly matched majority species of Sect.Pseudoptilagrostis. S. macroglossa and S. stapfii of the Sect. Stipa showed close phylogenetic relationship with Sect. Leiostipa taxa.2. MP tree from ITS sequences was similar with the chloroplast gene sequences phylogenetic trees. The Stipa species were divided into two well-defined monophyletic groups. Sect. Smironvia, Sect. Stipa and Sect. Leiostipa clustered into one clade, and Sect.Pseudoptilagrostis species clustered into another group.3. Both chloroplast gene sequences and ITS sequences analysis revealed that Stipa bungeana Trin., S. capillacea Keng and S. przewalskyi Roshev. had a far phylogenetic relationship with Sect. Leiostipa, so these three species should be defined as two independent sections and named as Sect. Bungeana and Sect. Przewlsyiana respectively.4. rbcL, trnL-F and ITS sequences analysis demonstrated that four species of Sect. Barbatae species (S. purpurea Griseb., S. breviflora Griseb., S. orientalis Trin. and S. roborowskyi Roshev.) were not clustered into a monophyletic group. However, it presented a complex phylogenetic relationship that S. orientalis and S. roborowskyi Roshev. clustered into one clade with Sect. Pseudoptilagrostis Tzvel.. All species of Sect. Smironvi species clustered into a well-defined monophyletic group.5. In the regional scales, Stipa species have a high level of genetic diversity. Eight primers of ISSR were applied with Stipa species that generated 301 reliable bands with the total number of 296 polymorphic bands, and the PPB was 98.30%. The Nei’s index and Shannon’s index were highest in Sect. Barbatae and lowest in Stipa przewalsky respectively. What’s more, the interspecific diversity (DST= 0.2760) was greater than intraspecific diversity (Hs), and Stipa had a high value of GST (0.8296) due to the abundant variation among species. AMOVA analysis confirmed that a significant part of genetic differentiation (85.51%) in Stipa was attributed to inter-populations variation. The genetic differentiation within sections (14.47%) is even lower than that within populations (15.6%).6. Genetic diversity order of each group of Stipa is:Sect. Barbatae Junge>Sect. P seudoptilagrostis Tzvel.> Sect. Smirnovia Tzvel.> Sect. Leiostipa Dum.> Sect. Bungeana> Sect. Przewalskyiana Z.Y.Chuo Sect. Leiostipa has the highest geneti c consistency, with an average value of 0.9615, followed by Sect. Barbatae, with a n average value of 0.9585, and average value of genetic consistency in Sect. Pseudop tilagrostis is 0.9366. The high degree of similarity within each group of Stipa is relate d to inter-specific hybridization between groups of Stipa species.7. The genetic diversity of Sect. Leiostipa was significantly correlated with longitude and altitude, and the genetic diversity of Sect.Pseudoptilagrostis was significantly related to annual precipitation, annual mean temperature, latitude and longitude. The cluster analysis based on ISSR and DNA sequences suggested that Stipa performed obvious regional differentiation. Its evolution is mainly carried out in two directions, one is adaption to the arid climate, and the other is adaption to the high altitude cold climate. Sect. Leiostipa, Sect. Stipa, Sect. Barbatae and Sect. Smironvia belong to the former, while Sect.Pseudoptilagrostis belongs to the latter.8. Small scale human disturbance altered the genetic diversity and genetic pattern. We performed inter-simple sequence repeat (ISSR) molecular marker analysis on populations of Stipa grandis and S. krylovii which were exposed to five consecutive years of varying degrees of grazing disturbance. The amplification results showed that the genetic diversity of both S. grandis and S. krylovii populations varied under different grazing intensities; the highest diversity (Nei’s index and Shannon’s index) were under moderate disturbance, whereas the lowest under the heavy grazing. The coefficient of gene differentiation (GST) of S. grandis and S. krylovii populations were 16.82%and 21.00%, respectively. AMOVA analysis confirmed that a significant part of the genetic differentiation in S. grandis and S. krylovii populations was attributed to the grazing effects. These results suggest that Stipa species were extremely sensitive to environmental changes. Spatial heterogeneity is an important cause of the regional differentiation and genetic variations in Stipa. |
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