Phylogenomics Of The Genus Corylus And Genetic Differentiation Of C.heterophylla Fisch.species Complex

Hazelnut belongs to the genus Corylus L.in the family Betulaceae,with approximately 20species(varieties)intercontinentally distributed in Asia,Europe and North America.Since the mid-19th century,scholars both at home and abroad have been engaged in the phylogenetic study of Corylus,but no definite conclusion has been reached so far,especially the origin,divergence history,and geographical distribution pattem of the genus have not been studied yet.China is the diversity center of Corylus species,of which the C.heterophylla Fisch.complex is the most widely distributed taxon,inclduing three closely related shrub species(C.heterophylla,C.kweichowensis,and C.yunnanensis).Their natural populations distribute continuously from the northeast to southwest of China and contain excellent genetic resources,but the studies on the species differentiation,population structure and genealogical relations are extremely limited,thus hindering the processes of resource conservation and selective breeding.In this study,Corylus species and populations of C.heterophylla complex were detailed sampled.Through 2b-RAD reduced-representation genome sequencing,the phylogeny and biogeography of Corylus were comprehensively studied at the nuclear genome level.Based on the whole chloroplast genome sequencing,the variation and evolutionary features of Corylus chloroplast genome were revealed,and phylogenetic reconstruction was conducted at the plastome level.Appling SCN loci as molecular markers,the genetic structure and species differentiation within C.heterophylla species complex were clarified using the methods of population genetics.The main results are as follows:1.Revealing the variation and evolutionary characteristics of the chloroplast(cp)genome of Corylus.The chloroplast(cp)genome of Corylus was a typical quadripartite structure with the genome size ranging from 159,822 to 160,358 bp.Within each cp genome,a total of 131genes were encoded,of which 18 genes were duplicated in two IR regions.The variation degree of LSC and SSC regions was significantly higher than that of IR regions,from which nine hotspot regions were selected as potential molecular markers.The IR/SSC boundary positions and their flanking genes of most species were relatively stable,while the IR/LSC boundary positions had different degrees of variation,especially the IR expansion of C.heterophylla,C.avellana,C.chinensis,and C.ferox var.thibetica.The cp genomes of Corylus contained abundant repeat structure which were mainly distributed in the intergenic sequences of LSC region.Chloroplast phylogenies supported the division of Betulaceae into two subfamilies(Betuloideae and Coryloideae),and also confirmed the monophyletic position of Corylus in the Coryloideae.The mechanism of chloroplast capture lead to the lineage relationships to be closely associated with geographic distribution rather than taxonomic relationships.2.The phylogenetic framework of Corylus was constructed,and the origin,evolution and migration routes were clarified.Based on genome-wide SNPs developed from 2b-RAD sequencing,phylogenies,species trees,and STRUCTURE analysis revealed roughly identical clustering relations in spite of certain differnces.The NO-RECOMBINATION phylogeny revealed the optimal taxonomic and phylogenetic relationships by dividing Corylus into four major clades.STRUCTURE inference and Neighbour-Net analysis uncovered extensive signals of introgressive hybridization and reticulate evolution among Corylus species,which were also the main cause of nucleo-cytoplasmic phylogenetic conflict.C.wangii probably originated from the sympatric hybridization of C.yunnanensis and C.ferox,whereas the ancestors of C.chinensis came from certain parents of Clade C and Clade D.The recent common ancestors of Corylus occurred in Southwest China during the middle Eocene and spread to Europe and North America through long distance dispersal,while climate fluctuation and habitat fragmentation since the Miocene led to the species diversification in East Asia.3.The population structure and genetic differentiation of the C.heterophylla species complex were analyzed.All the three species within the C.heterophylla complex showed high levels of nucleotide polymorphisms,of which C.kweichowensis was slightly higher than C.heterophylla and C.yunnanensis.The overall recombination levels of the three species at 10nuclear loci were consistent,with a mean R_mof 5.Neutral tests showed that most single-copy nuclear loci were consistent with neutral evolutionary hypotheses,with a few sites deviating significantly from neutral.At the species complex level,the genetic variation mainly came from within populations,followed by among populations within species,and finally among species.At the species level,the genetic differentiation within populations was significantly greater than that of among populations.The genetic differentiation between C.heterophylla and C.kweichowensis was not obvious due to their close relationship,and C.yunnanensis was the first to separate from C.heterophylla complex.A large number of high-frequency haplotypes were shared among the three species,and the medium-frequency haplotypes were shared by populations with overlapping regions of C.heterophylla and C.kweichowensis,C.kweichowensis and C.yunnanensis,while these unique haplotypes were only owned by single population or shared among few adjacent populations.In summary,this study deeply dissected the genetic background of the worldwide species of the genus Corylus and the Chinese C.heterophylla complex.These results lay a foundation for the germplasm innovation and molecular assisted breeding of Corylus in the future.