Abstract: The genetic diversity (allele frequency, mean heterozygosity, mean polymorphic information content(PIC), and Nei genetic distances) of preserved population of 11 native chicken breeds was analyzed by 20 microsatellite markers with high polymorphism. The clustering dendrogram was obtained eventually based on Nei genetic distances of 20 microsatellite. The results showed that 176 alleles were detected of 20 microsatellite loci in 11 native chicken breeds, the average number of alleles of each loci was 8.8 and the range of allele frequencies was 0.013～0.838, and 11 native chicken breeds shared 45 alleles. The average heterozygosity of 20 microsatellite loci was 0.6800～0.7432, the heterozygosity of Zang chicken was the highest( 0.7432), and that of Langshang chicken was the lowest( 0.6800).The range of PIC of 20 microsatellite loci were 0.6329～0.7023, which is higher than 0.5 and highly polymorphic. By the results of UPGMA tree, four groups were formed from 11 chicken breeds, the first group was Silky, Chahua, Xianju, Zang and Xiaoshan chicken; the second group was Luyuan and Langshan chicken; the third group was Gushi, Fatty and Dagu chicken; and the fourth group was Henan game chicken. The effect of preserved breeds and the differences among generations can be tested by the file foundation of molecular markers, which contained allele frequency, mean heterzygosity, and mean PIC of 20 microsatellite loci in different generation populations.