| Populus × euramericana cv.’74/76’(hereafter,poplar 107)is a superior clonal variety of Section Aigeiros of Populus.It is widely planted in North China and Huanghuai and other regions.And it is the largest poplar variety planted in Hebei Province.Poplar 107 has been propagated asexually for more than 30 years since it was introduced into China in 1984.There were some differences in growth and morphological characters in different places.As a single genotype clonal variety,what are the reasons influencing the morphological differences of poplar 107,and whether there are epigenetic variations during long-term asexual reproduction?Clarifying these issues is of great significance to the promotion,application and management of poplar 107.In this study,epigenetic variation of poplar 107 was studied from transcriptome and DNA methylation.One-year-old poplar 107 seedings collected from 12 regions in China were cutting in the same environment,and their growth and morphology indexes were measured regularly.The leaves of poplar 107 from four regions with significant differences were selected for DNA methylation and transcriptome sequencing to explore the methylation pattern of poplar 107 genome and its mechanism of regulation on gene expression.The effects of abiotic and biotic factors on the growth traits and wood properties of poplar 107 were also studied.Adult trees of poplar 107 were collected from different areas in Hebei Plain,the growth traits were investigated,and the chemical and physical and mechanical properties of the wood were measured.The effects of environmental factors on the growth traits and wood properties of poplar 107 plantation in the Hebei Plain were studied by stepwise regression method.Meanwhile,bacterial community diversity in different parts(heartwood,sapwood,root and rhizosphere)of poplar 107 was evaluated by high-throughput sequencing technology to study the relationship between wood property,wet heartwood disease and bacterial community in different parts of poplar 107.The main results were as follows:(1)After cuttings of poplar 107 from 12 regions in China,the difference of oneyear-old seeding height was not significant,and the ground diameter,seeding stem color and leaf color showed abundant variation.Among the 12 lines,the ground diameter of Xianyang(XY)of Shanxi province was significantly larger than that of the tissue culture seedlings(ZP)of our research group.The leaf color chroma b*value of XY was the largest,which was significantly larger than that of Mancheng(MC)of Hebei province and Taian(TA)of Shandong province.Four lines with obvious differences were selected,namely MC,TA,XY and ZP.The growth traits of two-yearold seedlings of the four lines showed obvious variation.The plant height and diamete at breast height of XY were always significantly higher than those of TA.The plant height of XY was significantly higher than that of MC in August and September,and the diamete at breast height of XY was significantly higher than that of ZP in July and August.(2)Transcriptome analysis was conducted on MC,TA,ZP and XY leaves by using transcriptome sequencing technology.The number of differentially expressed genes in ZP with XY comparison group was the largest,followed by MC and TA with XY.Compared with XY,the number of up-regulated genes of MC,TA and ZP was lower than that of down-regulated genes.Compared with XY,the differentially expressed genes of MC,TA and ZP comparison groups were mainly enriched in metabolic pathways and biosynthesis of secondary metabolites.Compared with the other three lines,metabolic pathways,biosynthesis of secondary metabolites,starch and sucrose metabolism,pyruvate metabolism and propionate metabolism,which were closely related to growth and development,were significantly expressed in XY.The differentially expressed key regulatory genes involved in carotenoid biosynthesis,flavonoid biosynthesis,flavone and flavonol biosynthesis affect leaf color.(3)Whole-genome methylation sequencing analysis of MC,TA,ZP and XY leaves was performed using sulfite sequencing technology,CG methylation was the main methylation form in the genome of poplar 107,with methylation levels ranging from 33.46%to 34.03%.CHG methylation followed,with methylation levels ranging from 17.03%to 17.73%.Most methylation sites were located in gene and intergenic regions.Most CG and CHG sites were hypermethylated state;most of the CHH sites were hemimethylated or hypomethylated state,which were relatively unstable.In the gene region,CG type had the highest methylation level in the genebody region,while CHG and CHH type had the lowest methylation level in the genebody region.In the transposon region,the methylation level of the three types showed a similar trend,that was,the methylation level was highest in the genebody regions.(4)In the upstream 2 kb region of the gene region,the CHH methylation level of XY was significantly lower than that of ZP.In the genebody region,the upstream and downstream 2 kb region of the transposon region,the CHH methylation level of XY was significantly lower than that of MC,TA and ZP.In CG and CHG methylation patterns,TA with ZP had the largest number of differentially methylated regions,followed by MC and XY with ZP.In CHH methylation pattern,MC with XY had the largest number of differentially methylated regions,followed by MC and TA with ZP.KEGG enrichment analysis showed that in the CG and CHG types,genes related to differential methylated regions in the six comparison groups were mostly enriched in metabolic pathways such as the biosynthesis of secondary metabolites,biosynthesis of amino acids,carbon metabolism,plant-pathogen interactions,and ribosome.In the CHH type,almost no genes related to differential methylated regions were enriched in metabolic pathways in the six comparison groups.(5)DNA methylation and transcriptome association analysis showed that found the influence of methylation level and methylation type on gene expression depended on gene location.Spearman correlation analysis of DNA methylation level and gene expression level in gene region showed that only a small part of genebody region had high positive correlation between methylation level and its expression level.In CG and CHH types,compared with XY,common genes of differentially methylated regionrelated genes and differentially expressed genes of MC,TA and ZP comparison groups were mainly involved in the growth and development of 107 poplars.In CHH type,there was no enrichment pathway for all common genes.(6)There were abundant variations in growth and wood properties of 25 poplar 107 from the Hebei Plain.The coefficients of variation(CVs)of mean annual increment of height,diameter at breast height,and volume were 14.47%,10.57%,and 22.40%,respectively.The CVs of cellulose,hemicellulose,and lignin content of the heartwood were 4.28%,10.71%,and 11.12%,respectively.The CVs of cellulose,hemicellulose,and lignin content of the sapwood were 3.24%,8.64%,and 8.32%,respectively.The CVs of cold water,hot water,and 1%NaOH extractive content in the heartwood were 21.64%,30.20%,and 10.57%,respectively.The CVs of cold water,hot water,and 1%NaOH extractive content in the sapwood were 27.51%,22.35%,and 7.79%,respectively.The differential shrinkage of the heartwood was classified as Grade 1,while that of the sapwood was classified as Grade 2.The air-dried density of both the heartwood and the sapwood were classified as Grade 2.The bending strength and bending elastic modulus of the heartwood and sapwood were also classified as Grade 2.The compressive strength parallel to grain were classified as Grade 3 for both the heartwood and the sapwood.Mean annual relative humidity,mean annual sunshine hours,soil pH and total K were the key factors affecting growth traits of poplar 107.Soil available P and total P were the key factors affecting the chemical properties of poplar 107 heartwood.Soil available P and alkali-hydrolyzed N were the key factors affecting the chemical properties of sapwood.Soil pH,total K and mean annual ground temperature were the key factors affecting the physical and mechanical properties of poplar 107 heartwood.Soil organic matter,pH and mean annual precipitation were the key factors affecting physical and mechanical properties of sapwood.(7)There were differences in the bacterial community diversity of poplar 107 heartwood in the northern,central and southern regions of the Hebei Plain,but no significant differences in bacterial community diversity among sapwood,root,and rhizosphere.The main environmental factor affecting the community structure of rhizosphere bacterial was mean annual ground temperature.The main environmental factors affecting the community structure of endophytic bacterial of poplar 107 were soil pH,total N,available K,temperature and sunshine duration.The heartwood,sapwood,root and rhizosphere of poplar 107 provided different ecological niches for bacterial communities.Relatively speaking,the community structure of rhizosphere bacteria was the most similar to that of root endophytic bacteria,and the less similar to that of sapwood endophytic bacteria.(8)A total of 16,283,473 high-quality reads were obtained from 16S rDNA highthroughput sequencing of 100 poplar 107 samples,and the number of OTUs after clustering was 6592,belonging to 41 phyla and 917 genus.The dominant bacteria of heartwood samples were Bacteroidetes(35.63%),Firmicutes(27.38%)and Proteobacteria(11.55%).The dominant bacteria of sapwood samples were Cyanobacteria(80.03%),Proteobacteria(15.56%)and Bacteroidetes(1.40%).The dominant bacteria of root samples were Proteobacteria(51.20%),Cyanobacteria(19.89%)and Actinobacteria(8.65%).The dominant bacteria of rhizosphere samples were Proteobacteria(34.73%),Actinobacteria(16.19%)and Acidobacteria(13.37%).(9)Compared with the heartwood of normal trees,the wood color of wet heartwood of poplar 107 was reddish brown.Wet heartwood disease reduced the wood quality of poplar 107.The diversity index(Shannon and Simpson)of endophytic bacteria in the heartwood of poplar 107 wetwood trees(WW)was significantly lower than that in the heartwood of normal trees(NW)(p<0.05),and the community structure between the two types of trees was significantly different.No significant differences in the alpha diversity index or community structure were observed between the sapwood,root tissue,and rhizosphere bacteria of the two types of trees(p<0.05).Proteiniphilum,Actinotalea and Methanobacterium were the dominant genera in WW-H.Proteiniphilum,Dysgonomonas and Bacteroides were the dominant genera in NW-H.The relative abundances of Proteiniphilum,Actinotalea,and Methanobacterium were significantly higher in the heartwood of wetwood trees than those of normal trees,and were positively correlated with the rate of wet heartwood.These three bacteria might be the main pathogens causing poplar wetwood disease.Compared with the heartwood bacterial community of WW,the heartwood bacterial community of NW had a more complex network,indicating that the disorder of the endophytic bacterial community in the heartwood of poplar 107 led to the occurrence of wet heartwood disease. |
