| Dalbergia odorifera T.Chen(Fabaceae),indigenous to Hainan Island,is a precious rosewood(Hainan hualimu)in China.However,only limited genomic information is available,which has resulted in a lack of molecular markers,limiting the development and utilization of the germplasm.In this study,we aim to enrich genomic information of D.odorifera,and develop a series of transferable simple sequence repeat(SSR)markers for Dalbergia species.Based on these newly developed SSR marekers,a comprehensive survey on genetic diversity was performed individually for the resource of D.odorifera,Dalbergia cochinchinensis Pierre ex Laness,and Dalbergia tonkinensis Prain.(1)Transcriptome sequencing was performed for D.odorifera by pooling leaf tissues from three trees.A dataset of 138,516,418 reads was identified and assembled into 115,292 unigenes.Moreover,35,774 simple sequence repeats(SSRs)were identified as potential SSR markers.A set of 19 SSR markers was successfully transferred across species of D.odorifera,D.tonkinensis,and D.cochinchinensis.In total,112 alleles(3-13 alleles/locus)were presented among 60 Dalbergia trees,and polymorphic information content(PIC)ranged from 0.38 to 0.75.The mean observed(Ho)and mean expected heterozygosity(He)was 0.34 and 0.40 in D.odorifera,0.27 and 0.32 in D.tonkinensis,and 0.29 and 0.33 in D.cochinchinensis,respectively.The cluster analysis classified these 60 trees into three major groups according to the three Dalbergia species based on the genetic similarity coefficients,indicating these newly developed transferable markers can be used to explore the relationships among Dalbergia species and assist genetic research.(2)We evaluated the genetic diversity of 42 wild D.odorifera trees from seven populations covering its whole native distribution Hainan Island.In total,19 SSR(simple sequence repeat)markers harbored 54 alleles across the 42 individuals,and the medium genetic diversity level was inferred by Nei's gene diversity(GD 0.36),observed(0.28)and expected heterozygosity(0.37).Among the seven wild populations,the expected heterozygosity(He)varied from 0.31(HNQS)to 0.40(HNCJ).The analysis of molecular variance(AMOVA)showed that only 3%genetic variation existed among populations.Moderate population differentiations among the investigated populations were indicated by pairwise Fst(0.042-0.115).Structure analysis suggested two clusters for the 42 individuals.Moreover,the seven populations were clearly distinguished into two clusters from both the principle coordinate analysis(PCoA)and neighbor-joining(NJ)analysis.Populations from Haikou city(HNHK),Baisha autonomous county(HNBS),Ledong autonomous county(HNLD),and Dongfang city(HNDF)were comprised cluster I,while cluster II comprised the populations from Wenchang city and Sanmengpo city(HNQS),Changjiang autonomous county(HNCJ),and Wuzhisan city(HNWZS).The findings of this study provide a preliminary genetic basis for the conservation,management,and restoration of this endemic species.(3)We used 19 SSR markers to assess the genetic diversity for 226 trees,including one in situ conservation population HN(42 wild trees)and two ex situ populations CM(96 seedlings)and JJ(88 grafting seedlings).Totally,the SSR markers harbored 75 alleles across the 226 D.odorifera individuals.Medium genetic diversity levels exhibited separately for these populations indicated by unbiased expected heterozygosity(uHe)of 0.38 in CM,0.37 in JJ,and 0.37 in HN,respectively.No significant difference was detected in these genetic diversity indices among these populations by the paired samples T tests,and minor differentiations showed among them inferred by the result of AMOVA(1%).Structure analyses were performed individually for the investigated populations which were clearly showed four,three,and two clusters for CM,JJ,and HN,respectively.AMOVA results showed 13%,11%,and 10%genetic variation existed among subpopulations in CM,JJ,and HN,respectively.The results of this study provided a comprehensive assessment on the genetic diversity and population structure for the large D.odorifera germplasm under conservation,and will be useful for the efficient and sustainable utilization of these genetic resources.(4)We collected a database of 251 trees(including 42 wilds and 210 cultivated individuals)across the whole southern China,evaluated genetic diversity and population structure of the database using 19 SSR markers,and develop a core collection for improvement and breeding programs.Totally,the markers harbored 77 alleles across the 251 individuals with PIC ranging from 0.03 to 0.66.Medium genetic diversity level was inferred by GD(0.38),I(Shannon's information index 0.65),Ho(0.33)and He(0.38).Structure analysis grouped the whole germplasm into four main reconstructed populations(RPs)and an admixture(MIX)by the model-based Bayesian procedure.Among the five subpopulations,the He varied from 0.30(RP3)to 0.38(RP4).Results of AMOVA showed 11%genetic variation existed among subpopulations,and moderate subpopulation differentiations among them exhibited with pairwise Fst ranging from 0.031 to 0.095.Moreover,a core collection of 31 D.odorifera including 6 wild and 25 cultivated individuals was developed,which was only 12.4%of the whole germplasm but conserved the whole genetic diversity.The results of this study provided additional insight for the genetic structure in the large D.odorifera germplasm,and the core collection will be useful for the efficient and sustainable utilization of the genetic resources,as well as efficient improvement in breeding programs.(5)Finally,genetic diversity of 70 D.cochinchinensis and 67 D.tonkinensis individuals were seperately evaluated using the 19 transferablilty SSR markers.The PPB(the percentage of polymorphic loci)was 68.42%and 100%across the D.cochinchinensis and D.tonkinensis,respectively.The germplasm of D.cochinchinensis,collected from Taihland and Combodia,exhibited medium genetic diversity and genetic variation level inferred by Ho(0.28),uHe(0.37),and Fst(0.173),respectively.The germplasm of D.tonkinensis,collected from Vietnam also showed medium genetic diversity level indicated by Ho and uHe of 0.24 and 0.32,respctively.Moreover,relatively high genetic variation level was inferred by Fst of 0.696.The results of this study provide a useful basis for the efficient and sustainable utilization of D.cochinchinensis and D.tonkinensis genetic resources.All the results provide useful information for the sustainable utilization and breeding programs on genetic resource of the Dalbergia species. |
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