Abstract: 【Objective】 The purpose of this study is to explore the genetic diversity and genetic differentiation of Yemili aboriginal chickens in the hope of providing theoretical basis for the breeding of this breed. 【Methods】 In this study, 15 microsatellite loci were used to analyze the genetic diversity of four groups (G₁:1 generatons, G₂:2 generatons, M₁:1 generatons, and M₂:2 generatons). 【Results】 The results showed that a total of 102 alleles were detected in 15 microsatellite loci in 120 individuals, and the average number of alleles (Na) per locus was 6.8.The average expected heterozygosity (He) and observed heterozygosity (Ho) values for the points were 0.708 and 0.660, respectively.The average polymorphic information content (PIC) (0.661) was highly polymorphism.The mean intra-population inbreeding coefficient (Fis), total population inbreeding coefficient (Fit) and inter-population genetic differentiation coefficient (Fst) were 0.051, 0.070 and 0.020, respectively, indicating that the low degree of inbreeding and genetic differentiation were not obvious.The population gene flow MCW0216 site was the largest (29.689), the smallest LEI0166 site (6.192), the average gene flow (Nm) was 15.091, and a high degree of gene communication among the four populations.Cluster analysis showed that four groups of Yemili aboriginal chickens were divided into two groups according to genetic distance. 【Conclusion】 In conclusion, Yemili aboriginal chickens have great selection potential and rich genetic diversity.