Four non-coding region, trnH-psbA 、 trnS-trnG spacer + intron 、 trnT-5'trnL and 5'trnL-trnF of 242 germplasm accessions of Malus sieversii from 3 sources of Gongliu, Huocheng and Xinyuan of Xinjiang Uygur Autonomous Region of China were amplified by four primers. Based on the genetic variation of the four chloroplast intergenic regions, the genetic variation and evolution of Malus sieversii populations were explored from the perspective of maternal inheritance. The results showed that the length of the four non-coding regions of chloroplast DNA was 3812 bp after sequencing, splicing, alignment and merging, and 171 variable sites were detected, including 6 singleton variable sites, 16 parsimony informative sites and 149 insertion-deletion gaps. Among the 242 accessions of Malus sieversii the number of variable sites of the regions trnH-psbA, trnS-trnG spacer + intron, trnT-5'trnL and 5'trnL-trnF were 68, 25, 77 and 1. The number of haplotypes for the four regions were 36, 6, 6 and 2, and after the four regions merged the haplotypes of chloroplast DNA fragments were 52. The region with the highest nucleotide and haplotype diversity was trnH-psbA (Hd=0.773 ， Pi=0.01982), and the nucleotide and haplotype diversity of 5'trnL-trnF was the lowest (Hd=0.025 ， Pi=0.00002). The cpDNA diversity of Malus sieversii with the four chloroplast DNA regions merged was high (Hd=0.806，Pi=0.00291). Tajima’s test showed that all the Tajima’s D values are statistically significant at P＜0.05, indicating that the overall variation of the four chloroplast regions had not followed the neutral theory of molecular evolution, and the pressure of natural selection is the main driving force of genetic evolution of Malus sieversii. The genetic variation of Malus sieversii mainly existed within populations. The distance between populations from Gongliu and Xinyuan of Xinjiang was closer than that between populations from Huocheng and Gongliu or from Huocheng and Xinyuan of Xinjiang. The genetic differentiation correlated with the geographical distances. Malus sieversii from the 3 sources were experiencing genetic differentiation, but also having frequent gene exchanges among different populations, and had a tendency to evolve to Xinyuan population in Xinjiang.