| Vicia is an important crop for green manure and high quality forage.It can also be used as food and medicine,and has high economic value.This study 9 wild and cultivated Vicia forage germplasm resources as the material,ITS morphological differences,not only more leaves,seeds and pollen morphological structure,also,by using the method of ITS molecular markers from two levels of phenotypic diversity and ITS sequence polymorphism on the genetic diversity research,the phenotypic clustering and molecular clustering results were analyzed.To clarify the genetic diversity of Vicia herbage resources,and to provide a certain theoretical basis for revealing the population relationship of Vicia herbage resources.The main results are as follows:(1)The coefficient of variation of 14 macro characters in leaves ranged from0.00% to 49.13%,among which the coefficient of variation of leaflet-apex serration was the smallest,while the stem pilose density was the largest.The genetic diversity index ranged from 0.00 to 1.06,among which the index of leaflet-apex serration was the smallest and the stem pilose density was the largest.The variation coefficient of leaf epidermis micromorphological characteristics ranged from 0.00% to 80.83%,among which type of stomatal apparatus had the smallest variation and stomatal density had the largest variation.The genetic diversity index ranged from 0.00 to 1.89,among which the cell shape index was the smallest and size of stomatal index was the largest.The cell shapes of 9 materials were irregular,and the stomatal apparatus types were irregular.There were significant differences in the other leaf epidermal micromorphology(P < 0.05),and the leaf morphological variation of the 9 Vicia was rich.(2)The variation coefficient of seed quality traits ranged from 23.39% to47.14%,with cotyledon color having the smallest variation and umbilical color having the largest variation.The genetic diversity index ranged from 0.53 to 1.15,among which the cotyledon color index was the smallest,and the testa color index was the largest.The variation coefficient of quantitative traits ranged from 5.28% to63.88%,among which the variation of seed length/ seed width was the smallest,the variation of weight of 100 seeds was the largest,and the genetic diversity index was between 1.06 and 1.68,among which the index of radicle length/radicle width was the smallest,and the index of the length of seed,the thickness of seed and umbilical length/seed length were the largest.(3)The pollens of the 9 materials were all single grains with oblate and spherical shapes,and there were obvious differences in the size of pollen grains.The pollens were oblate in the equatorial plane view,and trifid round and triangular in the polar plane view.There were two types of outer wall ornamentation: reticular ornamentation and brain ornamentation.The coefficient of variation of pollen traits ranged from 0.00 to 34.40%,among which the variation of the equatorial view was the smallest,and the variation of the ornamentationwas the largest.The genetic diversity index ranged from 0.00 to 1.68,where the index of the equatorial view and the aperture were the smallest,and the index of the polar axis length and the equatorial axis length were the largest.(4)Principal component analysis of 40 morphological indicators showed that high load in the seven principal component index for leaf size,hundred seed weight,seed length,seed width,the length of umbilical,umbilical color,relative leaflet size,shape of the lobule base,radicle tip shape,stomata width,stomata length,umbilical length/seed length,seed length/seed width,lower leaflet position,stipple shape and umbilical shap,16 indicators,the above indicators revealed the main reasons for the phenotypic differences of the 9 Vicia in this study.The results of cluster analysis showed that when the relative distance was 20,it could be divided into two groups.The first group was divided into two subgroups,with a total of 8materials.Materials 2,3,4,5,6 and 7 constituted the first subgroup,materials 8 and 9were included in the second subgroup,and material 1 alone constituted the second group.(5)The results of ITS sequence analysis showed that the 5.8S region of r DNA was relatively conserved,with 72 mutation sites in ITS1 region and 37 mutation sites in ITS2 region,respectively.The variation rate of ITS1 region was 29.6% higher than that of ITS2 region,17.5%,and the haplotype diversity was 0.972 ± 0.064.The nucleotide diversity was 0.032±0.004,the genetic distance of K2 P was 0.0044～0.5616,and the percentage of polymorphism was 94.64%.The NJ tree showed that the 9 Vicia materials and Pisum sativum L.were divided into three categories,and the outgroup— — Pisum sativum L.was located at the base of the tree.The first material,the four-seeded Vicia,had a distant relationship with the other materials and formed a branch,while the other 8 materials were grouped into one category.(6)In terms of leaf,seed and pollen morphology,the evolution of Vicia sepium L.,Vicia cracca L.and Vicia sativa var.Sativa was relatively primitive,while the evolution of Vicia tetrasperma(L.)Schreb.,Vicia villosa Roth var.and Vicia villosa Roth(hairy vetch)was relatively high.(7)The results of morphological clustering and phylogenetic tree based on ITS sequence showed that the positions of all the materials in the phylogeny were basically the same.The taxonomic status and genetic relationship of the 9 test materials corroborate each other at the morphological and molecular levels,obtaining relatively complete and systematic genetic information,and clarifying the genetic basis of germplasm resources. |
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