ISSR Analysis Of Genetic Diversity Of Robinia Pseudoacacia Populations From China

Robinia pseudocacia is drivered from the Appalachian Mt and the Ozank Mt in the east of America. The planting area of it is the second largest in the world fast-growing broad leaf thees, which is only smaller than that of Eucalyptus. It is widely introduced by many countries for its strong adaption, fast growth and various use. Together with Poplar and Eucalyptus, Robinia pseudocacia is known as one of the most successfully introduced trees in the world. It was in 1898 when it was introduced into China and now it is widely planted all over the country. During a long process selection by the nature and human, the populations of Robinia pseudocacia contain abundant types of gene and have formed genetic diversity. The objective of the study is to evaluate the genetic diversity and genetic structure within and among populations of Robinia pseudoacacia from China detected by AFLP in order to discuss the genetic variation and evolvement of its breeding resources. The main results from this study are as follows:1. Extraction genome from Robinia pseudoacacia.1. Factors influencing ISSR analysis, including the purification and concentration of DNA, concentration of Mse I /EcoR I and T4DNA ligase and time for reaction, ingriedents for solution were studied during the AFLP process .An optimized system of silver staining was established for Robinia pseudoacacia AFLP analysis. Results indicate that the usage of DNA template is 200-400ng, Mse I /EcoR I is3U,T₄DNA ligase is 1.5U, digestion time is 2h and ligation time is 12h, the products of ligation are diluted to 10 times for preamplification and the products of preamplification are diluted to 10 times for selective amplification. It was shown that fingerprinting map of Robinia pseudoacacia can be constructed with primer E+3 /M+3 and that there are more amplified bands with high polymorphism and good quality.2. The genetic diversity of 10 Robinia pseudoacacia provenances collected from China were analyzed by AFLP technique. Totally 100 individuals were selected and analyzed by amplifications using 10 pairs of AFLP primers screened, each primer pair produced 38⁹7 bands and 752 polymorphic bands were obtained, of which 352 (46.8%) were polymorphic. As for Robinia pseudoacacia at species level, The average number of alleles per loci (Na) of the 10 populations was 1.4987, and the effective number of alleles per loci (Ne) was 1.2328, the proportation of polymorphic loci(P) was 49.87%, the mean Nei's gene diversity index (H) was 0.1403, the Shannon's information index (I) was 0.2160, indicating that there is some genetic diversity of this weed. As for Robinia pseudoacacia, the coefficient of gene differentiation among populations within species (Gst) was 0.3390, indicating that in the 10 Robinia pseudoacacia Provenances, the genetic differentiation among the populations was relatively less , whereas within the populations was relatively greater.3. Groups cluster analysis based on the UPGMA method indicated that although the 10 populations could be divided into 3 big groups. The first group included Yangling, Jingchuan, Chifeng, Weifang and Changli, whose latitude ranged from 34.27°N to 42.28°N. The second group included populations of Feixi, Xinzhou, Lushi and Kunming, whose latitude ranged from 25.04°N to 34.06°N. The latitude of all populations of the second group was lower than the first group. The third group only contained Chaoyang, whose latitude was 41.58°N. From the UPGMA analysis, we could see that there were some correlation between the genetic relationship among populations and the geographic distribution, while evident geographic variation patterns of the Robinia pseudocacia seed sources had not been formed.4. The space spreading structure of species' genetic variation has an important affection to make a scientific tactic of protection and use of the gene resources. In order to adapt to the difference of the environments of populations, each population formed its own specially. For example, the third group, only population of Chaoyang, whose index of the genetic diversity was obviously higher than that of the first two groups. The percentage of the specific bands of population of Yangling was the highest, which had the genotypes that the other populations did not have. This was possibily related to the huge introduction of the overseas Robinia pseudocacia in Yangling. So it was first to protect groups and individuals within the group, especially those grown in the different geographic structure and had the genotypes that the other populations did not have. The genetic diversity of Robinia pseudocacia from China mainly existed within the populations, so the initial selections in the breeding programs should concentrate on intrapopulational variation in improvement of Robinia pseudocacia. The genetic improvements should mainly concentrate on the individuals' improvement within populations though selection of the superior trees, while the effect of the populations' selection was much small.