Study On The Population Genetic Diversity And Altitude Adaptability Of Rhododendron Rubiginosum In Southwest Plateau Mountains Of China

Rhododendron rubiginosum Franch.is widely distributed in northwest to northeast Yunnan,southwestern Sichuan and southeast Xizang of China.The species always distributed in forest gaps between the elevations of 2,500 and 4,200 m,and are the dominant species in the community.Thus,it is an important component of the southwestern plateau mountain ecosystem with high ecological and economic values.Up to now,the studies of R rubiginosum mainly focus on the chemical composition and hermotolerance,no research about the population structure,genetic diversity,genetic variation,multi-omics and the adaptation of this species were conducted.Thus,in the present study,we studied the genetic diversity,genetic structure and demographic history of R.rubiginosum populations using microsatellite markers and DNA fragment（cp DNA:mat K,psb A-trn H and rbc L;n DNA:ITS and RPC＿1）.Furthermore,we also conducted a comparative analysis of global transcriptome and gene expression pattern of R rubiginosum at low-,middle-and high-elevation gradients（Rr L,Rr M and Rr H）.The results of this paper will not only provide a theoretical basis for the protection,resource assessment and sustainable development and utilization of R rubiginosum,but also provide a reference for exploring the response mechanism of mountain plants in the southwest plateau to environmental changes.The major results and conclusions are summarized as follows:1.Population genetics analysis of R.rubiginosumBased on the six polymorphic microsatellite loci and five DNA fragments（rbc L,mat K,psb A-trn H,ITS and RPC＿1）sequence and analysis,we assessed the levels of genetic diversity,population structure and demographic history of R rubiginosum.The results indicated that the species have high levels of genetic diversity（N_a=7.771、N_e=4.645、H_e=0.680、PPB=100%）,and mediate levels of genetic differentiation(0.05<F_ST=0.1345<0.15)at the species level based on SSRs.AMOVA analysis showed that genetic variation mainly distributed within population,accounting for 86.06%.The inbreeding coefficient F_is=0.106>0 based on SSRs indicated that there is a certain degree of inbreeding within population of R.rubiginosum.The species has high historieal and low contemporary gene flow between 8 populations.In addition,PCo A,UPGMA clustering and Structure analysis showed that the 8 populations first clustered into two branches and then differentiated into 3 groups,the closer geographical groups were firstly clustered;Mantel test showed that the genetic distance and geographic distance exists a significant correlationship（R²=0.266,P<0.05）.Cp DNA and n DNA sequence and analysis found a significant genetic differentiation（H_T>H_S）,and a significant phylogeographic pattern(N_ST>G_ST)among the populations of R.rubiginosum.The bottleneck effect test showed that the populations had no bottleneck effect in recent years.Additionally,the neutrality tests and mismatch distribution analysis of cp DNA and n DNA revealed that R.rubiginosum did not experienced a recent population expansion.2.Transcriptomics and altitude adaptability of R.rubiginosumBased on Illumina Hi Seq4000 sequencing,we obtained the transcriptome data from low（Rr L,2950 m）,middle（Rr M,3550 m）,and high（Rr H,3550 m）altitude distribution of R.rubiginosum,respectively（accession number:SUB53668370）.A total of 396,446 unigenes and 65,802 potential SSRs were identified in R.rubiginosum.Comparative transcriptome analysis from the three elevation gradients revealed that 846differentially expressed unigenes（DEGs）,with 421 up-regulated and 425down-regulated in Rr L vs.Rr M.Meanwhile,7,777 DEGs,with 4,059 up-regulated and3,718 down-regulated in Rr L vs.Rr H.Through enrichment analysis of the GO and KEGG pathways,441 DEGs were significantly enriched into pathways related to plant photosynthesis,carbohydrates,hormones,and secondary metabolism.Among these DEGs,102 DEGs were related to photosynthesis,15 of them showed up-regulated expression,and 87 were down-regulated.172 DEGs related to the carbohydrate were identified,89 of them showed up-regulated expression,and 84 were down-regulated.We identified 39 DEGs associated with the metabolic synthesis of hormones,among these DEGs,28 and 11 DEGs showed up-regulation and down-regulation.A total of 190DEGs involved in secondary metabolite synthesis were identified in R.rubiginosum leaves,117 of them showed up-regulated expression,and 73 were down-regulated from low to high elevation according to the global expression profiles of R.rubiginosum.Thus,based on the above results,we conclude that R.rubiginosum adapts to the heterogeneous environmental conditions under different altitude gradients by regulating the gene expression related to its photosynthesis,carbohydrates,plant hormones and secondary metabolism.