Salix viminalis L. is an important shrub that has potential for use as a bioenergy crop, for phytoremediation of heavy metal contaminated soil and sewage sludge treatment. It is mainly distributed in the northeast of China, but the species has not yet been used a resource here. We examined the genetic diversity and population structure of populations from the Ergun basin and West Liao basin using 20 microsatellite markers. A high level of genetic diversity (Na = 16.45, He = 0.742) was detected for S. viminalis, and populations from the Ergun basin exhibited higher genetic diversity and private alleles numbers than the West Liao basin. The 12 populations could be divided into two clusters by both Bayesian analysis and UPGMA clustering which were consistent with the populations derived from the two basins. Moderate population differentiation (FST = 0.076) was shown in S. viminalis, and AMOVA analysis confirmed that most of the genetic variation (86.13%) was attributed to individual differences within populations, while 11.49% was attributed to differences between basins and 2.38% to differences within each basin. Significant correlations of FST/(1–FST) with log (geographic distance) among 12 populations (r = 0.634, p < 0.00) and 10 populations within the Ergun basin (r = 0.482, p = 0.0002) indicated that geographical distance was the principal factor influencing genetic structure. As most of genetic variation exist within populations, so protection measures should be focused on populations with higher genetic diversity and unique alleles, such as Tuli, Mordaga downstream, Zhadun1 and Genhe.