Microsatillite Development And Genetic Diversity Of Agrilus Mali

The wild apple, Malus sieversii(Ldb.) Roem forests are the precious germplasm gene pool of apples, and have been listed as the protected species and endangered plants of China. With the invasion of Agrilus mali Matsumura to Xinjiang, the wild apple forests have been seriously damaged by this pest. Many wild apples died because of the damaging of Agrilus mali. On the other hand, A. mali is damaing the cultivated apple trees, causing economic lost of the local people. In the current study, the magnetic beads enrichment method was used to develop seven polymorphic microsatellite loci for A. mali. Both the microsatellites obtained and the mitochondrial COI gene was used to analyzed the genetic diversity of seven A. mali population from wild apple trees in Xinjiang Province and one population from Liaoning Province. The results are important for the further investigation of the population expansion mechanism and development of integrative management strategies of the pest in wild apple forests. The main results are as follows:1. The bead enrichment method was used to develop Agrilus mali microsatellites. A total of 248 clones containing microsatellite were acquired, with the positive clone rate of 82.6%. In addition AC and AG repeats used in the probes, the CT, ATCA and TCGT repeats were also obtained. The repeat times of the microsatellites ranged from 16 to 20. Among the 248 microsatellites, 131 ones were perfect microsatellites, accounting for 52.8%; 90 ones were imperfect, accounting for 36.3%; 27 ones were mixed, accounting for 10.9%. Primers were designed to amplify the micosatellite loci. Seven polymorphic loci with stable amplification reaction systems were obtained. Basing on the analysis of the obtained seven microsatellite loci, 102 alleles were found in samples from eight geographic populations. The number of alleles was from 10 to 23, the effective alleles number were between 1.6738 and 12.2179, the observed heterozygosity was between 0.3125 and 0.9621, the expected heterozygosity was between 0.4041 and 0.9239, the polymorphic information content was between 0.3036 and 0.7961.2. Basing on the analysis of the obtained seven microsatellite loci, the genetic distances of seven populations from the Xinjiang Province ranged from 0.0714 to 0.5206. The genetic distances among populations from Xinjiang Province and Liaoning Provicne werebetween 0.4041 and 0.7876. The genetic similarities among populations from Xinjiang Province were between 0.5942 and 0.9311. The genetic similarities among populations from Xinjiang Province and Liaoning Province were between 0.4549 and 0.6676. The genetic distances and genetic similarities between population from Xinjiang were significant lower than that between population from Xinjiang and Liaoning according to t test, with t=2.056,df=26, P=0.00014, and t=2.056,df=26,P=0.00033, respectively.3. The mitochondrial COI gene was used to analyze the genetic diversity of Agrilus mali samples from Xinjiang and Liaoning. A total of 19 variable sites and 10 haplotypes were found in the sequences obtained. The Hap1 haplotype were shared by seven populations from Xinjiang. No population expansion and continued growth patterns were found for the population analyzed. There was suignificant genetic diffence between populations from Xinjiang and Liaoning.