Forty-two microsatellite loci were analysed in 23 highly inbred chicken lines derived from Leghorn, Jungle Fowl, Fayoumi and Spanish breeds. Line-specific alleles among breeds and lines were detected. The band-sharing (BS) values were calculated and the proportion of shared alleles distances (Dps) were estimated. The BS values and Dps between sets of MHC-congenic lines ranged from 0·74 to 0·96, and 0·05–0·35, respectively. The BS values between each pair of noncongenic Leghorn lines were 0·32–0·97, and between Leghorn and exotic (Jungle Fowl, Fayoumi and Spanish) breeds were 0·03–0·55. The Dps between Fayoumi lines and other lines were much larger (0·66–1·34) than within Leghorns, and the Jungle Fowl breed had the largest distances with other lines (1·12–5·38). The phylogenetic consensus tree that was constructed grouped these 23 inbred chicken lines into four different clusters. These results are in accordance with the origin and breeding history of these inbred lines, which indicates that the use of microsatellites for the study of genetic biodiversity is accurate and reliable. In addition, the significance and value of inbred chicken lines in molecular genetic research is discussed.