Seed Morphological Variation Of Different Sources And Molecular Identification Of Cultivars/Clones Of Sophora Japonica L. By SRAP Marker

Sophora japonica L. is a tree species with timber, medicinal, edible and ornamental values. This species is naturally distributed in China, Korea and Japan, and widely planted in many parts of the world.Seed morphology was examined in this study. 130 families in 7 populations of S. japonica were sampled. Seed length, seed width, the ratio of seed width to length, seed surface area, seed volume and 1000-seed's weight were measured in order to find out genetic variations and variation patterns. It was expected that research results would provide scientific bases for the assessment at genetic resources and strategy for conservation and utilizition of S. japonica. Phenotypic variation among and within populations were analyzed by ANOVA and correlation analysis. Genetic materials of varieties and clones of S. japonica were used for DNA extraction and the best SRAP-PCR reaction system selection for further development of the SRAP analysis. It was intended in this study that a theoretical basis to show genetic relationships at molecular level.The main findings and conclusions are as follows:1 Genetic variations were, to a certain extent, found in phenotypic properties of seed, coefficient of variation was ranged from 5.24% to 14.57%. There were significant differences in seed morphological characteristics among and within populations. The average differentiation coefficient among populations was 53.11%, which was higher than that within population. No significant correlation was found in geographic variations of seed with latitude and longitude. It was indicated in cluster analysis that 7 populations of S. japonica were divided into 3 groups:1) Taian groups; 2) Hubei and Beijing groups; 3) Zhejiang, Xi'an, Xianyang and Henan populations. It was found that there was no any significant geographical variations in seed morpholgy, which suggested that introduction and cultivation in wide range of area or discontinuous sampling would be responsible for the phenomenon. 2 Orthogonal experimental design was applied to establish the optimal reaction system for SRAP-PCR (20 uL): Taq polymerase 1.5 U, dNTPs 0.30 mmol / L, the template DNA 90 ng, Mg²⁺2.0 mmol / L, primer 0.3 umol / L, ddH₂O fill to 20 uL.3 There were 12 primer combinations that PCR product bands were clear, highly polymorphic selected from 120 primer combinations, which were applied in genetic relationship and molecular identification study of varieties and clones of S. japonica. The rang of PCR amplification of 12 primer combinations are mostly concentrated in bands between 100bp-2000bp. Of the total of 430 bands amplified, 415 were polymorphic loci. The percentage of polymorphic fragments were 96.60%, of which 4 pairs of primers polymorphism were 100%. It was showed in the results that multiple genetic loci of varieties and clones could be detected with SRAP markers. Primers combination Em9-Me12 could be used to distinguish different varieties of S. japonica. Rich and clear SRAP fingerprints were also obtained.4 Levels of genetic similarity in coefficients (SM) were calculated and a dendrogram was constructed by the unweighted pair group method of arithmetic average (UPGMA). SM value was in the range of 0.68-0.89, with an average similarity coefficient of 0.73, indicating the similarity between varieties and clones of S. japonica were relatively high, which means the genetic base was relatively narrow. Based on the similarity coefficients of 0.76, varieties and clones were clustered into four categories. The second category was consisted of 4 and 35 of S. japonica; the fourth category inclued S. japonica var. pendula Loud. (LZ), S. japonica'Caozhou 2'(C2) and S. japonica f. flavi-rameus (JY). The rests were clustered into the first category, which could be divided into two subgroups. S. japonica var.oligophylla Franch. itself was clustered in the third category, which indicated that there were large differences in different shapes of leaves.