In the present study, 10 short tandem repeats (STR) (HPRTB, DXS6789, DXS6799, DXS6804, DXS7130, DXS7132, DXS7133, DXS7423, DXS7424, and DXS8378) on X chromosome were investigated among four ethnic populations (431 individuals) in northwest of China, in order to learn about the genetic diversity, forensic suitability, and possible genetic affinities of the populations. The polymorphism information content (PIC) values for the ten loci range from 0.340 to 0.810, and the observed heterozygosity values for the ten loci range from 0.397 to 0.860, the results indicated that the ten loci have a moderate degree of variability in the four populations, and each population has its own characteristics of genetic structure. A Neighbor-joining (NJ) tree constructed on the basis of the generated data shows very low genetic distance between Han, Mongolian and Tibetan, Dongxiang and Tu as well as between Salar and Bonan populations. Our results based on genetic distance analysis are consistent with the results of earlier studies based on linguistics and the immigration history and origin of these populations. These STR loci on the X chromosome studied here are not only useful in showing significant genetic variation between the populations, but also are suitable for human identity testing among the four populations.