The allele frequencies, heterozygosity (H), mean heterozygosity, polymorphism information content (PIC) and genetic relationships were studied in Luyuan chicken, Gushi chicken, Tibetan chicken, Baier chicken, Xianju chicken, Chahua chicken, Dagu chicken,Beijing Fatty chicken, Langshan chicken, Henan Game chicken,Taihe Silkies chicken and Xiaoshan chicken using seven microsatellite DNA markers. The results showed that there were differences among allele fequencies of seven microsatellite loci in 12 Chinese native chicken breeds. Among the 12 Chinese native chicken breeds, mean heterozygosity of Luyuan chicken was the highest (0.5929), and that of Chahua chicken was the lowest (0.3514). There were similar results in PIC. By fuzzy cluster analysis was found that the genetic relationship between Taihe Silkies chicken and Henan Game chicken was the nearest, and that between Gushi chicken and the other chicken breeds was the most distant. The 12 Chinese native chicken breeds were divided into three group by fuzzy cluster analysis. The first group included Taihe Silkies chicken, Henan Game chicken, Langshan chicken, Xiaoshan chicken, Dagu chicken, Beijing Fatty chicken and Luyuan chicken, the second group included Chahua chicken, Tibetan chicken, Xianju chicken and Baier chicken, and the third group included Gushi chicken.