The Update Of Genetic Linkage Map And Qtls Mapping Of Important Traits In Eucommia Ulmoides

Eucommia ulmoides is an important economic forest species,which has application values of medicine and rubber.Some progress has been made in the selection of superior varieties of E.ulmoides,which greatly promoted the development of E.ulmoides industry.However,traditional breeding often takes decades to fully evaluate and release new cultivars,which restricted the progress of genetic improvement of E.ulmoides.Marker-assisted selection can obtain individuals with large gain or directly obtain superior varieties and clones through molecular marker or genetic engineering,which has greatly improved the selection efficiency and breeding accuracy.In this study,the hybrid F₁population?152 individuals,‘Xiaoye'בQinzhong No.1'?were employed as the materials,and a large number of SSR primers were developed from transcriptome sequencing to optimize the original genetic linkage map in E.ulmoides.The growth,phenology,secondary metabolite content and other traits of F₁ population were monitored for 10 consecutive years.Through multi-years QTLs mapping and candidate genes searching,the QTLs/genes expressed continuously and stably were identified.The system of molecular marker-assisted selection in E.ulmoides was constructed and used for the selection of superior trees in F₁population,which provides theoretical basis for the implementation of E.ulmoides juvenile screening and multi-traits collaborative improvement.It has important guiding significance to accelerate the process of directional improvement and breeding in E.ulmoides.The main results are as follows:1. Transcriptome analysis and development of SSR primers in E.ulmoides.Based on transcriptomic data,1,870 pairs of effective SSR primers?producing clear bands similar to the expected band size?and 351 pairs of SSR primers with high stability and polymorphism were developed.By transcript profiling of leaves and fruits in E.ulmoides,65 candidate genes of terpenoid biosynthesis pathway were identified.Through comparative transcriptome analysis,29 differentially expressed genes related to terpenoid biosynthesis were screened out.2. The update of genetic linkage map of E.ulmoides.Combined with the literature and transcriptome screening results,365 pairs of SSR primers were used for PCR amplification with the mapping population and two parents,and 456 polymorphism markers were obtained.Among them,22?4.8%?markers showed segregation distortion?p?0.05?.The updated genetic linkage map of E.ulmoides consisted of 19 linkage groups with 869segregating markers covering 1913.29 c M.The number of mapped markers per linkage group ranged from 15 to 151.The map size of the linkage groups ranged from 59.13 c M to172.21 c M.The mean genetic distance between adjacent markers was 2.20 c M.The estimated length of E.ulmoides genome was 2,015.05 c M.In this study,the constructed linkage map covered 94.95%of the estimated genome length.3. The 22 phenotypic traits of E.ulmoides F₁ hybrid population for five consecutive years?2015-2019?were statistically analyzed.All the traits were obviously separated in the mapping population and normally distributed.11 traits?tree height,breast diameter,ground diameter,number of leaf vein,petiole length,chlorogenic acid content,rutin content,Eu-rubber content,leaf length,leaf width and leaf area?have shown?extremely?significant correlation among five consecutive years.The remaining 11 traits?crown diameter,leaf length-width ratio,weight of single leaf,branch length,branch diameter,branching number,branching angle,number of leaves,total weight of dry leaves,date of bud burst and date of leaf exhibition?showed no correlation among more than 3 years.There was a certain degree of positive correlation between different traits.However,there was a significant negative correlation between branching angle and branch length,branch diameter,branching number,number of leaves and total weight of dry leaves.The content of Eu-rubber in leaves was negatively correlated with the content of chlorogenic acid and rutin.4. Observe and analyze the growth patterns of 22 phenotypic traits in F₁ hybrid population of E.ulmoides?152 individuals?for 10 consecutive years?2010?2019?.The growth of each trait have increased slowly with years.In the seedling stage?2a?,the leaf spreading area was large,the color was dark and the leaves were thick,which help to improve the photosynthesis of seedlings and store more organic matter and energy.At a relatively low age?1a?5a?,E.ulmoides was in a physiological stage that was conducive to the accumulation of secondary metabolites?chlorogenic acid,rutin,and Eu-rubber?.The date of bud burst and leaf exhibition were sensitive to the external environmental factors?temperature?.The growth of branch length,branch diameter,leaf number and total weight of dry leaves have increased significantly in 8a?2017?,and the trees began to bear fruit,suggesting that the rapid growth period of E.ulmoides began from 8a which indicated that the plant reached the maturity stage.5. QTLs mapping for important traits in E.ulmoides.A total of 432 QTLs were identified for 22 traits among 10 years.The LOD values of QTLs ranged from 3.01 to 37.52,among which 155 have reached the genome-wide LOD threshold value,accounting for 35.9%of the total number of QTLs.204 Q TLs have shown significantly correlated with traits through Kruskal-Wallis analyses,accounting for 47.2%of the total number.The phenotypic contribution of each QTL was 9.0%?85.8%.Except for QTLs of branching number and leaf area,the other QTLs have identified overlapping among different years,but no Q TLs at the same position were detected in consecutive ten years.Q TLs identified for different traits also have overlap interval.QTLs related to muti-traits and muti-years were speculated to be the key factor affecting E.ulmoides growth and development.6. The prediction of candidate genes in Q TL intervals.According to the transcriptome sequencing results,43 sequences located in the Q TLs interval were obtained,and 19sequences were located at the peak of the LO D value which were presumed to be closely related to the target traits.Through BLASTX and gene annotation,a total of 28 sequences?candidate genes?were identified with 27 annotation information,while the gene function of the remaining 15 sequences were unknown.The functional annotations of the candidate genes were divided into five categories:cellular structural components?3?,intracellular functions?8?,plant tissue formation?4?,signal transduction and defense responses?8?,and metabolism?4?.There were 2 sequences identified to be related with terpenoid pathway.7. The molecular marker-assisted selective breeding in E.ulmoides.Four molecular markers with high detection efficiency?EQ457-200a?em1me6-170?UBC886-2200 and em4me7-550?were selected.Based on molecular marker-assisted selection and phenotypic data,12,7,9,9 and 5 individuals were selected for high content of Eu-rubber,high content of medicine,high economy,high growth and comprehensive type,respectively.The suitable tree age for juvenile selection of E.ulmoides was 5a.