Evolution Of Whole Chloroplast Genome,Phylogeny And Whole-genome Duplication Of Gossypium

Cotton genus contains eight diploid genome groups(A-G and K)and one single allotetraploid group AD.The Gossypium contains about ~53 species,46 diploid and 7allotetraploid.However,up to now,the phylogeny and polyploid origin of Gossypium have been controversial.In this study,we used Illumina sequencing determine the chloroplast genome sequence of 16 species of cotton genus and the transcriptome sequences of 37 species,and combined with the data released from the database to analyze the differentiation and molecular evolution of Gossypium species.To identify the structural variation and sequence evolution of the chloroplast genome,phylogeny and the whole-genome duplication of Gossypium genus,and reveal the species differentiation and evolution mechanism inside the cotton genus.The main results of this study are as follows:1.The 40 chloroplast genome sequences of 33 cotton genus species were compared and analyzed,the result show that the chloroplast genomes of Gossypium species were highly conserved,of which sizes range from 159,039 bp to 160,433 bp;These genomes had a quadripartite molecule structure(LSC,SSC and a pair of IRs).In total,2860 long sequence repeats(>10 bp in length)were identified,where the F-genome specie had the largest number of repeats(G.longicalyx : 108)and E-genome specie had the lowest(G.stocksii : 53).There is a significant difference in the level of variation in different regions of the chloroplast genome of Gossypium,and the non-coding regions had greater variation than coding regions.We identified 10 divergence hotspot regions,i.e.,rpl33-rps18,psbZ-trnG(GCC),rps4-trnT(UGU),trnL(UAG)-rpl32,trnE(UUC)-trnT(GGU),atpE,ndhI,rps2,ycf1,and ndhF,which could be useful molecular genetic markers for future population genetics and phylogenetic research.Selective stress analysis of protein-coding genes and found 10 positive selection genes associated with the adaptive evolution of cotton species(atpB,atpE,rps2,rps3,petB,petD,ccsA,cemA,ycf1 and rbcL).Phylogenetic analysis showed that the cotton species were mainly divided into six evolutionary branches,similar to the previous studies,but the relationship between the diploid C,G,and K genomic species is still confusing,nested in a large evolutionary branch.Combined with the geographical distribution of cotton genus,it is speculated that it may be the recent radiation differentiation between species,incomplete lineage screeningand introgression/hybridization between different lineages.2.Upland cotton and sea island cotton are widely cultivated polyploid cotton,and their semi-wild species have excellent characteristics such as high quality and multi-resistance,and have close genetic relationship with cultivars.In order to further comprehensively analyze the origin and evolution history of cotton genus,a total of 10 species of chloroplast genomes were newly sequenced in semi-wild species of upland cotton,sea-island cotton and a K-genome specie.Based on 78 chloroplast genome protein coding genes of cotton genus,a phylogenetic tree was constructed.It was found that the cotton species were still divided into six previously identified genetic branches,and all 13 D genome species aggregated into one higher support.The upland cotton and its seven semi-wild species have a close genetic relationship,of which the relationship between G.hirsutum race latifolium and G.hirsutum is the closest,revealing that cultivated upland cotton may originate from the G.hirsutum race latifolium species.In addition,all the allotetraploid species are clustered into one,and the A genome species are closely related to the AD allotetraploid species.The results confirmed that the maternal donor of allotetraploid species probably belonged to A genome species.The time of differentiation test found that the main diploid branch of cotton was about 7.1 million years ago(Mya),similar to the results of previous studies.3.The phylogenetic tree was constructed using single-copy nuclear genes which were identified among 37 cotton species transcriptome data and 4 Gossypium genome data.The results showed that the three genomic species of C,G and K genome formed a monophyletic clade,which was basically consistent with the traditionally classified karyotype.The inconsistent developmental relationship between the chloroplast genome and the single-copy nuclear gene of Gossypium species may be caused by the recent frequent hybridization/introgression and the evolutionary rate and different genetic characteristics of differentiation between different gene markers..4.Whole-genome duplication(WGD)is the core of eukaryotic evolution and an it is the important evolutionary force of plant diversity.In this study,transcriptome sequencing and de novo assembly were performed on 37 Gossypium species,yielding 51,803,618-116,210,674 clean reads,assembling 41,480-78,402 unigenes.Using the distribution ofparalogous genes,two WGDs were detected in Gossypium species.The ancientr doubling event occurred in the early Pliocene,about 31.88-38.25 Mya;the more recent doubling event occurred in 2.0-6.59 Mya,WGDs may accelerate the formation and diversification of Gossypium species.