Phylogeography And Population Structure Of Endemic Walnut (Juglans Sigillata) In Southwest China Based On CpDNA And EST-SSR

Southwestern（SW）China is an area of active tectonism and erosion,yielding a dynamic,deeply eroded landscape that influences the genetic structure of the resident populations of plants and animals.Iron walnut（Juglans sigillata Dode）is an endemic,deciduous tree species native to this region of China and cultivated there for its edible nuts.We sampled 36 wild Iron walnut populations from locations throughout the species’range in SW China and genotyped a total of 765 individuals at 5 chloroplast DNA regions and 22 nuclear microsatellite loci.The data was used to infer levels of genetic differentiation,genetic structure,and contemporary/historical levels of gene flow.The results and conclusions of this study are as follows:1.Genetic diversity of Iron walnut.Based on cpDNA markers,Iron walnut showed relatively low genetic diversity（H_d=0.2026;π=0.00080）.Based on EST-SSR markers,the expected heterozygosity（H_E）of the 36 populations was lower than that of other species of the genus juglans（J.nigra:H_E=0.807;J.mandshurica:H_E=0.77）.The reason for this may be that Iron walnut is an endemic species with relatively small population sizes,and because of genetic drift,the founder effect,and other random causes,often have lower genetic diversity than more common and widely distributed species.2.Genetic differentiation of Iron walnut.Based on cpDNA markers of maternal inheritance,the level of genetic differentiation of the population was low(G_ST=0.194).The most likely reason for this is the recent large population fragmentation and repeated gene flow.Based on EST-SSR marker data,F_STT value relative to other temperate wind pollination tree is higher.In southwest China,the terrain is complex and climate is multiterminal changing.The walnut maintained a fragmented spatial structure throughout the quaternary period,leading to the increase of F_ST.AMOVA results showed that the genetic differentiation was higher within the population,which may be affected by the heterogamy system of Iron walnut.3.The genetic structure and gene flow of Iron walnut.Based on EST-SSR data analysis,the genetic structure of Iron walnut showed that the northern and southern assemblages of the distribution range of species were clustered into two groups.It was found that when STRUCTURE result K=2,there were unbalanced historical and contemporary gene flows from Cluster B to Cluster A.4.The evolutionary history and dynamic range of the Iron walnut.Based on Chloroplast DNA（cpDNA）and EST-SSR markers,Iron walnut were divided during the pliocene-pleistocene period.We used bayesian and reductive methods for phylogenetic analysis.Iron walnut were divided into two distinct branches.The differentiation time of branch A and branch B was2.65 Ma.ABC simulation analysis showed that the internal differentiation time of walnut was1.34 Ma.The uplift of Yunnan-Guizhou plateau in southwest China may have occurred in late Miocene-pliocene,resulting in habitat fragmentation due to global cooling and intensified Asian monsoon.The current geographic distribution of Iron walnut reveals three scattered refuges in the Yunnan-Guizhou plateau:Hengduan mountain region,Wuliang mountain region,and one refuge in the northeastern part of the species distribution.Species distribution area simulation shows that Iron walnut experienced a range of contraction from the last interglacial period to the last ice age,Iron walnut retreated to about 25°N,at the same time,the bayesian skyline results show that the effective population size experience contraction in0.25 Ma,then experience slight expansion,this phenomenon is the main reason is the LGM period is more and more cold and dry,after the climate warming.In a word,Species distribution models were produced to predict the evolution and historical biogeography of Iron walnut,and to estimate the impacts of climate oscillations and orographic environments on the species’demography.Our results indicated that J.sigillata had relatively low genetic diversity,high inter-population genetic differentiation,and asymmetric inter-population gene flow.Results of ABC analyses indicated that the distinct south subpopulations and north subpopulations persisted in a long-term glacial refuge in southwestern China since the beginning of the last glacial period,whereas increasingly colder and arid climates at the onset of the last glacial maximum might have fostered the fragmentation within refugia populations of J.sigillata.Finally,we found that recent habitat fragmentation has led to a reduction of population connectivity and increased genetic differentiation by genetic drift in isolated populations.Our results support a conclusion that geological and climatic factors since the Miocene triggered the differentiation,evolutionary origin,and range shifts of J.sigillata in the studied region.