| Firmiana danxiaensis is a rare and endangered species endemic in China,which has special distribution characteristics and high ornamental value.In order to conservation this rare species,it is important to know the genetic diversity and population structre along whole distribution areas.In this study,we developed 16 polymorphic EST-SSR markers,based on the transcriptome sequences.Using these locs,the genetic structure of F.danxiaensis populations was analyzed.These genetic information were significance for the conservation of this speceis.The main results of the study as below.1.Development and characteristic of EST-SSR markes fox F.danxiaensisIlluminaHiSeqTM 2500 was used for transcriptome sequences.After removal of redundant reads,the short-reads assembly software Trinity was used for transcriptome de novo assembly.A total of 17 858 SSRs were detected from 79 920 unigenes,which distributed in 12 103 unigenes.The SSR incidence rate was 15.14%.Trinucleotide was the most motif repeat(7 794 SSRs sequcnes),accounting for 43.60%of the total repetitive type.88 EST-SSR loci were selected for primer synthesis,which the repeat number more than 9 times.Among these primes pairs,73 was successful amplification,and 16 loci has polymorphism tested by 16 individules.For each locus,the number of alleles(Na)varied from 3 to 10.The average polymorphism information content(PIC)was 0.361.Ten loci deviate from HWE equilibrium due to,heterozygote access.Six loci may exist null alleles.The results of PCA analysis showed that the developed EST-SSR loci have the ability to identify different population and even each individuals.These loci can be used for futher molecular genetic studies of F.danxiaensis.2.Genetic diversity and population structure of F.danxiaensis18 EST-SSR markers were used for the study of genetic diversity and population structure of F.danxiaensis.Total 131 alleles were detected from 9 populations(213 individuals).At the species level,F.danxiaensis has moderate-level genetic diversity,with the expected heterozygosity(He)was 0.536,and the average allelic richness(AR)was 3.642.The group of Danxia Mountain has higher genetic diversity than that of Nanxiong group,with the Shannon's Information Index data were 0.974 and 0.917,respectively.The results of AMOVA analysis showed that the genetic variation among individuals within the population was 79.66%(p<0.01),the genetic variation among the population within the group was 5.28%(p<0.01),and the genetic variation between the groups was 15.06%.(p<0.01).The genetic differentiation coefficient of Fst and Gst were 0.147 and 0.135,respectively,which suggested a strong genetic differentiation among the populations of F.danxiaensis,especially between two groups of Danxia Mountain and Nanxiong.Structure analysis demonstrated that the models with K=2 provided satisfactory explanations of the observed data.The bar plot for K=2,all populations were separated into two gene pools,which related with the laction of each populations(Danxia Mountain and Nanxiong).With the increasing of K(KF=3 and KF=4),new gene pools appeared in Danxia Mountain populations.Poptree system and PCA principal component analysis also showed consistent relationships anomg populations and groups.Mantel test showed that there was a significant positive correlation between genetic distance and geographic distance(r=0.8335 P=0.01).Bottleneck results suggest that some groups experienced recent bottleneck events.The DIYABC software was used to simulate the divergent event and estimate divergent time between two groups.A comparison of posterior probabilities of the four scenarios using local linear regression indicated that scenario 1 had the highest support with a probability of 0.4303.ABC analysis provided good support for scenario 1 with the same time divergence of Danxia Mountain and Nanxiong groups.The divergent time may 86 100?143 500 years ago,and the size of the effective population is 7 290 and 5 550 for two groups,respectively.The Cenozoic Quaternary crust tectonic movement and human disturbance were the main factors for the genetic structure of F.danxiaensis populations.3.The proposed protection strategyWith high genetic diversity and high private allele(Danxia Mountain),we are supposed to establish natural reserves and artificially manage the breeding of populations;For genetically low populations(Nanxiong),artificial conservation and expansion of inter-regional gene flow is necessary;Strengthen research on other aspects of wild populations,such as seed biology,physio-ecology,the breeding system and breeding mechanism of F danxiaensis,which have important guiding significance for the development of scientific and reasonable protective measures. |
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