Genetic Diversity Evaluation And Core Germplasm Construction Of Robinia Pseudoacacia L.germplasm Resources

Black locust(Robinia pseudoacacia L.)belongs to leguminous deciduous trees,which have important feeding and ecological value.In this study,a lot of black locust resources were collected from several regions in China.The genetic variation of these resources was systematically determined from phenotypic,physiological and molecular levels,revealing the genetic diversity of black locust.At the same time,the core collection of black locust resources was constructed by using various research strategies,which provided a basis for the effective utilization and protection of these plants.The major conclusions were described as follows:The results of investigation and analysis on 13 leaf traits of 1054 black locust samples showed that the total mean coefficient of variation of all traits was more than 10%,which showed that the black locust resources had abundant phenotypic variation,and the variation of leaflet circularity was the smallest and the highest stability at both the geographic population level and the clone level,while the trait of leaflet area was reversed.The mean phenotypic differentiation coefficient among geographic populations was 48.235%,indicating that the variation within geographic populations was the main source of phenotypic variation.There was no significant correlation between phenotypic traits and geographical distance at the geographic population level or clonal level(P>0.05).According to the measured phenotypic traits,different numbers of stress-resistant locusts were selected in each base.Three physiological traits of these black locust samples were measured,the results showed that these resources had rich physiological variation,and the variation coefficient of each trait is 10.558%?49.748%(at the clonal level)and 7.800%?40.649%(at the geographic population level),respectively,showing the smallest variation of chlorophyll content and the highest stability.The mean physiological differentiation coefficient among geographic populations was 39.143%,indicating that the variation within geographic populations was the main source of physiological variation.There was no significant correlation between physiological traits and geographical distance at the geographic population level or clonal level(P>0.05)According to the measured physiological traits,different numbers of stress-resistant locusts were selected in each base.EST-SSR molecular marker development,potential microsatellite repeats were detected from 36,533 unigenes and a total of 5,072 potential EST-SSR loci were identified.We successfully designed 2,486 pairs of primers were successfully designed,and primers from the same unigenes sequence were deleted,resulting in 1,697 pairs of SSRs.After strict control of the screening conditions,286 pairs of primers were selected.Furthermore,thirty-two samples of black locust and 9 species of leguminous plants were used to test the effectiveness of the amplification of this group of markers,and finally 45 pairs of EST-SSR markers were successfully developed.Subsequently,genetic diversity of locust germplasm resources was evaluated at molecular level.Forty-eight SSR markers has been selected from 91 pairs of SSR primers by polyacrylamide gel electrophoresis(PAGE).After the amplification of 1054 black locust germplasms by capillary electrophoresis,by examining the validity of the markers,35 pairs of SSR markers were finally determined for analysis of 367 samples of black locust geographical population(based on the origin).The results showed that 439 alleles were identified,with an average of 12.543 for each locus.The average polymorphism information content is greater than 0.5 with abundant polymorphism,and the mean observed(Ho)and expected(He)levels of heterozygosity were 0.495 and 0.553,respectively,indicative of a deficit of heterozygotes in black locust.The results of population genetic structure analysis divided 367 samples into two subpopulations,while the population structure was weak.AMOVA suggested that a majority of the genetic differentiation variation could be attributed to within-geographic-population differences.The genetic distance and identity results indicated that geographic distance was not a dominating factor influencing the distribution of black locust.Furthermore,after the validity test of 48 pairs of SSR primers,36 markers were determined to analyze the genetic diversity of 687 samples from black locust bases in China.The results showed that 587 alleles were identified,with an average of 16.306 for each locus.The average polymorphism information content(PIC)and gene polymorphism(H)were 0.061 and 0.645 respectively,indicating that each point had high polymorphism.The average expected heterozygosity(He=0.608)was higher than the observed heterozygosity(Ho=0.551),showing that a deficit of heterozygotes in black locust.The results of population genetic structure analysis divided 687 samples into four subpopulations,and a certain group structure has emerged.Mantel tests between geographic population and geographic distance showed geographic distance is not a dominant factor influencing the distribution of black locustConstruction of core germplasm of black locust.Firstly,molecular data were calculated by different methods.After comparison,it was determined that the core germplasm based on R software had the highest level of genetic diversity.Secondly,a small number of phenotypes and physiological individuals screened by M strategy in PowerCore software were selected as the required germplasm.Finally,we constructed a core population of 338 individuals.The core population and the original population constructed by comparison showed that the differences were not significant in genetic diversity,phenotype or physiological level,indicating that the constructed core group maintains the variation of the original group and can better represent the original group.