Study On Genetic Structure Of Osmanthus Serrulatus Natural Populations

As one of the rare spring flowering Osmanthus species in Oleaceae,Osmanthus serrulatus Rehd.usually known as Baoxing Osmanthus,which is endemic to China and usually distributed under the evergreen broad-leaved forests at elevations of 1,800-2,400 m.Since the characteristics of overyear seed germination,slowly natural regeneration and seriously human damaged,the wild populations are threatened critically in the local place and sporadically distributed in the medium to high altitude mountain slopes in the west of Sichuan Province,such as Emei Moutain,Xiling Snow Mountain and Dongla Moutain.The population in Dongla Moutain was the largest wild Osmanthus population of the world up to now.The detailed genetic diversity information of O.serrulatus were the most basic preconditions for the endangered mechanism researching,conservation strategy making and selective breeding,while the study on it was barely blank.Based on the comprehensive field investigation,there were 330 individuals collected from 13populations in the local place,and 18 pairs of SSR primers were selected to analyze the population genetic structure and genetic diversity.The study could provide molecular evidences for species variation and differentiation for O.serrulatus populations,and benefit for the effective protection and reasonable development on the species.The main conclusions are as follows:A total of 100 pairs were randomly selected from the 2435 primers obtained from transcriptome sequencing,and finally 15 pairs of high polymorphic and stabilized primers with length of 110 bp to 280 bp were screened and used for population genetic structure and genetic diversity analysis,accounting for 15%of the total number of random primers.There were 166 alleles detected by 15 pairs of screened primers and 3 pairs of universal primers at the species level.And the average number of alleles?N_T?and effective number of alleles?N_e?of O.serrulatus were 9.222 and 4.141 respectively,while the mean of Shannon diversity index?I?was 1.153,the mean of observed heterozygosity?H_o?was 0.218,the mean of expected heterozygosity?H_e?was 0.694 and the mean of Nei's diversity index was 0.838,which indicating the high genetic diversity.At the population level,the effective number of alleles of 13populations ranged from 3.309 to 4.477,and the range of observed heterozygosity?H_e?among populations was from 0.121 to 0.404,which were all lower than the expected heterozygosity.It indicated a relatively high level of genetic diversity and less hybrid in different natural populations.The genetic diversity of 13 wild populations from the highest to the lowest was:HY2>DY>LS2>HY3>HY1>BX2>EMS>JKH>BX3>DY2>BX4>BX1>LS1.The analysis showed that the populations located in the center of the natural distribution area of O.serrulatus had higher genetic diversity,which also showed that the level of population genetic diversity had a direct relationship with the strength of human disturbance.The coefficient of genetic differentiation(F_st)was 0.095 on average,which indicated that9.5%of the genetic variation occurred among populations and 90.05%occurred within populations.The gene flow?N_m?was 3.240,indicating a high gene exchange and a small genetic differentiation between the 13 populations.Bottleneck analysis showed that 4 populations?JKH?D1?HY1?LS1?experienced a recent bottleneck effect.Mantel test indicated that there was a certain correlation between the population genetic distance and geographical distance,with the r value of 0.15155.Analysis of UPGMA showed that the closer geographical groups in BaoXing were firstly clustered,which further confirmed the geographical isolation affected the genetic differentiation among populations.The STRUCTURE analysis results were slightly different from the UPGMA clustering results,and the 330 samples were divided into 4 groups and 41 individuals had a certain mixture of pedigrees,which further demonstrated the existence of gene exchange among populations.The result of principal component analysis?PCoA?was consistent with STRUCTURE analysis.The study results showed that the O.serrulatus maintained a relatively high level of genetic diversity and had a huge evolutionary potential,which indicating the genetic factors were not the main reason for the population decrease.It may be related to the dioecious and lack of seeds,the seed physiological maturation and difficult to germinate in natural conditions,the high environmental requirements and poor adaptation capacity,and the excessive human interference such as habitat destruction and over exploitation.Hence,the in situ and ex situ conservation strategy should be put into cation together,and given the proiority to protect the high genetic diversity populations.At the same time,it is necessary to increase publicity,raise awareness of protection and reduce human disturbance.