| Swida wilsoniana(S.wilsoniana)is a kind of deciduous tree with strong stress resistance and ecological adaptability.The high fruit oil content of S.wilsoniana has both edible and industrial value.At present,the improved variety breeding of S.wilsoniana mainly adopts the traditional methods,such as natural seed selection,crossing,and ploidy breeding.However,the traditional breeding has the disadvantages of long breeding cycle and low efficiency,which limits the breeding and popularization of the improved varieties.Molecular assisted breeding can shorten breeding cycle and improve breeding efficiency.Due to the lack of comprehensive genomic information and annotation,then development of molecular assisted breeding is impeded.In this study,based on natural distribution resources,germplasm resource and hybrid progeny resources,the whole genome of S.wilsoniana were sequenced and the characteristic of genome were systematically investigated.In order to thoroughly elucidate the genome of S.wilsoniana and the molecular genetic mechanism of important economic traits,the analysis of differentially expressed genes and key genes in the regulation of major reproductive traits in flowering stages,construction of genetic linkage maps andQTL mapping of related traits,and the establishment of phenotypic differentiation and genotype associations were conducted.The main results are listed as follows:1.The whole genome analysis of S.wilsoniana was carried out,and the whole genome information and annotations of S.wilsoniana were obtained,which enriched the genetic resources.The whole genome sequencing and preliminary assembly of S.wilsoniana were conducted using the third-generation PacBio sequencing technique.Results showed thatthe size of S.wilsoniana genome was about 904.65Mbp,among which the contig N50 and scaffold N50 were 4.24 Mbp and 78.01 Mbp,respectively.0.78%of heterozygosity and 55.22%ofrepeat sequences was observed for the genome of S.wilsoniana,whose splicing sequence wasanchored to 11 chromosomes by HiC technology.Results showed that 94.3%of eukaryotic core genes were identified in the genome of S.wilsoniana.Long terminal repeats(LTRs)class transposons are the main transposons,accounting for 48.83%of S.wilsoniana genome.A total of 31,640 structural genes were annotated with ach gene containing an average of 4.89 exons,of which 29,852 genes were annotated as protein coding genes.In addition,499.55 Mbp repeats and 3,618 noncoding RNAs were annotated.2.The historical evolutionary analysis of S.wilsoniana was carried out,which clarified the phylogenetic position of S.wilsoniana and identified two whole-genome duplication events.A total of 39,832 gene families were clustered by comparative genomic analysis of 15 species(S.wilsoniana,wild carrot,ginseng,ash,coffee,sunflower,soybean,walnut,Litsea Cubeba,hickory,sesame,Arabidopsis,oil palm,Jatropha,A.trichopoda),including 7,001 common gene families and 93 sharedsingle copy gene families.Compared with other species,there are 1,136 species-specific gene families(including 3,147genes).S.wilsoniana is closely related to wild carrot,ginseng,sunflower,ash tree,sesame,and coffee.And the evolution rate is fast and early,the time of species divergence is between 102.8×106～108.9×106 years,about 105.9×106 years ago.Compared with their recent ancestors,S.wilsoniana has significantly expanded by 89 gene families(including 1464 genes),significantly contracted by 28 gene families(including 89 genes),and 39 non-synonymous mutant genes were generated.S.wilsoniana experienced two genome-wide replication events,andtheywere 96×106 years,and 360 × 106 years ago.3.The synthesis and regulation analysis of important economic traits of S.wilsoniana was carried out,and functional genes and transcription factors related to reproductive growth traits were identified.Transcriptome analysis of S.at different growth stages shows that there are 307 differentially expressed genes in flower bud stage,early blooming stage and full blooming stage,and the number of up-regulated genes or down-regulated genes during flower bud stage to early blooming stage were higher than those during early blooming stage to full blooming stage.There are only two kinds of transcription factors SE and MXE in different growth stages of S.wilsoniana.From flower bud stage to early blooming stage,differentially expressed genes were enriched with a total of 115 KEGG metabolic pathways,among which carbon metabolism pathway and plant hormone signal transduction pathway were the dominant.From early blooming stage to full blooming stage,differentially expressed genes were enriched with 97 KEGG metabolic pathways,and the most enriched one was pentose and glucuronic acid conversion KEGG metabolic pathway.4.Using the hybridized F1 seedlings of S.wilsoniana as the mapping population,the genetic linkage map of S.wilsoniana was constructed by simplified genome sequencing(RAD-seq).Through SNP genotyping statistics,the genotype classification results obtained are hk×hk,lm×ll,nn×np,and the corresponding marker numbers are:8,707,25,287,23,095,a total of 57,089,and after correction,there are 8,707,25,277,23,087,a total of 57,071 loci.After filtration and screening,a total of 9,110 markers remained,accounting for 15.96%before filtering.The number of makers of genotype hk×hk,lm×ll,and nn×np were 2490,3353 and 3267,accounting for 28.59%,13.26%and 14.15%before filtration,respectively.The SNP marker of hh,hk,kk,nn,np,11,and lm genotype in F1 population were 39,942,79,814,39,866,101,124,105,619,111,453 and 101,382 respectively,accounting for 5.77%,11.52%,5.75%,14.60%,15.25%,16.09%,14.63%and 16.40%of the total genotyping respectively.5.The QTL mapping analysis of important economic traits of S.wilsoniana was carried out,and the genome-wide association was established according to phenotypic differentiation and genotype.QTL mapping for phenotypic traits of F1 offspring seedlings was conducted accordingtophenotypic differentiation and genetic variation detection of progeny populations.A total of 804 QTLs(LOD>3)were detected,which were distributed in 9 linkage groups,namely Lgl,Lg2,Lg3,Lg4,Lg5,Lg6,Lg7,Lg8 and Lg9,with 183,51,104,92,5,75,292,1 and 1 QTL markers,respectively.According to the variation of petiole length,leaf length,leaf width and leaf area at seedling stage,QTLs were accurately located and mapped on the genetic map.There were 61 QTLs related to parameter petiole length,leaf area,leaf length and leaf width,which were distributed on 9 linkage groups,forming 2 QTLs clusters.QTLs related to parameter petiole length,leaf width and leaf lengthwere clusteredwithin 94.40～107.54 cM on linkage group Lg3,while QTLs with parameter leaf width and leaf length were clusteredwithin78.09～79.06 cM on linkage group Lg7.The region of two QTLs clusters needs to be focused on in the subsequent mining of functional genes of growth-related traits.Nine significant correlation loci by phenotypic path and onesignificant locus by phenotypic seed settingwere screenedfrom parental resources.One significant locus was screened by phenotypic leaf length for progeny resources.No significant loci but 10 related loci were screenedfor petiole length,leaf width and leaf area.To sum up,above-mentioned results provided a theoretical basis for elucidating the genetic mechanism of important economic traits of S.wilsoniana,and afforded genetic information resources for further in-depth molecular genetics research in the future and identification of functional genes for many traits such as oil synthesis in S.wilsoniana. |
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